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Sample records for carrying mobilizable transgenes

  1. Transgenic pig carrying green fluorescent proteasomes

    PubMed Central

    Miles, Edward L.; O’Gorman, Chad; Zhao, Jianguo; Samuel, Melissa; Walters, Eric; Yi, Young-Joo; Prather, Randall S.; Wells, Kevin D.; Sutovsky, Peter

    2013-01-01

    Among its many functions, the ubiquitin–proteasome system regulates substrate-specific proteolysis during the cell cycle, apoptosis, and fertilization and in pathologies such as Alzheimer’s disease, cancer, and liver cirrhosis. Proteasomes are present in human and boar spermatozoa, but little is known about the interactions of proteasomal subunits with other sperm proteins or structures. We have created a transgenic boar with green fluorescent protein (GFP) tagged 20S proteasomal core subunit α-type 1 (PSMA1-GFP), hypothesizing that the PSMA1-GFP fusion protein will be incorporated into functional sperm proteasomes. Using direct epifluorescence imaging and indirect immunofluorescence detection, we have confirmed the presence of PSMA1-GFP in the sperm acrosome. Western blotting revealed a protein band corresponding to the predicted mass of PSMA1-GFP fusion protein (57 kDa) in transgenic spermatozoa. Transgenic boar fertility was confirmed by in vitro fertilization, resulting in transgenic blastocysts, and by mating, resulting in healthy transgenic offspring. Immunoprecipitation and proteomic analysis revealed that PSMA1-GFP copurifies with several acrosomal membrane-associated proteins (e.g., lactadherin/milk fat globule E8 and spermadhesin alanine-tryptophan-asparagine). The interaction of MFGE8 with PSMA1-GFP was confirmed through cross-immunoprecipitation. The identified proteasome-interacting proteins may regulate sperm proteasomal activity during fertilization or may be the substrates of proteasomal proteolysis during fertilization. Proteomic analysis also confirmed the interaction/coimmunoprecipitation of PSMA1-GFP with 13/14 proteasomal core subunits. These results demonstrate that the PSMA1-GFP was incorporated in the assembled sperm proteasomes. This mammal carrying green fluorescent proteasomes will be useful for studies of fertilization and wherever the ubiquitin–proteasome system plays a role in cellular function or pathology. PMID:23550158

  2. Plasmid pEC156, a Naturally Occurring Escherichia coli Genetic Element That Carries Genes of the EcoVIII Restriction-Modification System, Is Mobilizable among Enterobacteria

    PubMed Central

    Werbowy, Olesia; Kaczorowski, Tadeusz

    2016-01-01

    Type II restriction-modification systems are ubiquitous in prokaryotes. Some of them are present in naturally occurring plasmids, which may facilitate the spread of these systems in bacterial populations by horizontal gene transfer. However, little is known about the routes of their dissemination. As a model to study this, we have chosen an Escherichia coli natural plasmid pEC156 that carries the EcoVIII restriction modification system. The presence of this system as well as the cis-acting cer site involved in resolution of plasmid multimers determines the stable maintenance of pEC156 not only in Escherichia coli but also in other enterobacteria. We have shown that due to the presence of oriT-type F and oriT-type R64 loci it is possible to mobilize pEC156 by conjugative plasmids (F and R64, respectively). The highest mobilization frequency was observed when pEC156-derivatives were transferred between Escherichia coli strains, Enterobacter cloacae and Citrobacter freundii representing coliform bacteria. We found that a pEC156-derivative with a functional EcoVIII restriction-modification system was mobilized in enterobacteria at a frequency lower than a plasmid lacking this system. In addition, we found that bacteria that possess the EcoVIII restriction-modification system can efficiently release plasmid content to the environment. We have shown that E. coli cells can be naturally transformed with pEC156-derivatives, however, with low efficiency. The transformation protocol employed neither involved chemical agents (e.g. CaCl2) nor temperature shift which could induce plasmid DNA uptake. PMID:26848973

  3. Delivering Transgenic DNA Exceeding the Carrying Capacity of AAV Vectors.

    PubMed

    Hirsch, Matthew L; Wolf, Sonya J; Samulski, R J

    2016-01-01

    Gene delivery using recombinant adeno-associated virus (rAAV) has emerged to the forefront demonstrating safe and effective phenotypic correction of diverse diseases including hemophilia B and Leber's congenital amaurosis. In addition to rAAV's high efficiency of transduction and the capacity for long-term transgene expression, the safety profile of rAAV remains unsoiled in humans with no deleterious vector-related consequences observed thus far. Despite these favorable attributes, rAAV vectors have a major disadvantage preventing widespread therapeutic applications; as the AAV capsid is the smallest described to date, it cannot package "large" genomes. Currently, the packaging capacity of rAAV has yet to be definitively defined but is approximately 5 kb, which has served as a limitation for large gene transfer. There are two main approaches that have been developed to overcome this limitation, split AAV vectors, and fragment AAV (fAAV) genome reassembly (Hirsch et al., Mol Ther 18(1):6-8, 2010). Split rAAV vector applications were developed based upon the finding that rAAV genomes naturally concatemerize in the cell post-transduction and are substrates for enhanced homologous recombination (HR) (Hirsch et al., Mol Ther 18(1):6-8, 2010; Duan et al., J Virol 73(1):161-169, 1999; Duan et al., J Virol 72(11):8568-8577, 1998; Duan et al., Mol Ther 4(4):383-391, 2001; Halbert et al., Nat Biotechnol 20(7):697-701, 2002). This method involves "splitting" the large transgene into two separate vectors and upon co-transduction, intracellular large gene reconstruction via vector genome concatemerization occurs via HR or nonhomologous end joining (NHEJ). Within the split rAAV approaches there currently exist three strategies: overlapping, trans-splicing, and hybrid trans-splicing (Duan et al., Mol Ther 4(4):383-391, 2001; Halbert et al., Nat Biotechnol 20(7):697-701, 2002; Ghosh et al., Mol Ther 16(1):124-130, 2008; Ghosh et al., Mol Ther 15(4):750-755, 2007). The other major

  4. Delivering Transgenic DNA Exceeding the Carrying Capacity of AAV Vectors

    PubMed Central

    Hirsch, Matthew L.; Wolf, Sonya J.; Samulski, R.J.

    2016-01-01

    Gene delivery using recombinant adeno-associated virus (rAAV) has emerged to the forefront demonstrating safe and effective phenotypic correction of diverse diseases including hemophilia B and Leber’s congenital amaurosis. In addition to rAAV’s high efficiency of transduction and the capacity for long-term transgene expression, the safety profile of rAAV remains unsoiled in humans with no deleterious vector-related consequences observed thus far. Despite these favorable attributes, rAAV vectors have a major disadvantage preventing widespread therapeutic applications; as the AAV capsid is the smallest described to date, it cannot package “large” genomes. Currently, the packaging capacity of rAAV has yet to be definitively defined but is approximately 5 kb, which has served as a limitation for large gene transfer. There are two main approaches that have been developed to overcome this limitation, split AAV vectors, and fragment AAV (fAAV) genome reassembly (Hirsch et al., Mol Ther 18(1):6–8, 2010). Split rAAV vector applications were developed based upon the finding that rAAV genomes naturally concatemerize in the cell post-transduction and are substrates for enhanced homologous recombination (HR) (Hirsch et al., Mol Ther 18(1):6–8, 2010; Duan et al., J Virol 73(1):161–169, 1999; Duan et al., J Virol 72(11):8568–8577, 1998; Duan et al., Mol Ther 4(4):383–391, 2001; Halbert et al., Nat Biotechnol 20(7):697–701, 2002). This method involves “splitting” the large transgene into two separate vectors and upon co-transduction, intracellular large gene reconstruction via vector genome concatemerization occurs via HR or nonhomologous end joining (NHEJ). Within the split rAAV approaches there currently exist three strategies: overlapping, trans-splicing, and hybrid trans-splicing (Duan et al., Mol Ther 4(4):383–391, 2001; Halbert et al., Nat Biotechnol 20(7):697–701, 2002; Ghosh et al., Mol Ther 16(1):124–130, 2008; Ghosh et al., Mol Ther 15

  5. Neuropathological changes in transgenic mice carrying copies of a transcriptionally activated Mos protooncogene.

    PubMed

    Propst, F; Rosenberg, M P; Cork, L C; Kovatch, R M; Rauch, S; Westphal, H; Khillan, J; Schulz, N T; Vande Woude, G F; Neumann, P E; Newmann, P E

    1990-12-01

    Independent transgenic mouse lines carrying the mouse Mos protooncogene linked to a retroviral transcriptional control sequence display behavioral abnormalities including circling, head tilting, and head bobbing. This dominant phenotype shows various degrees of penetrance in different transgenic founder animals and lines. Neuronal and axonal degeneration, gliosis, and inflammatory infiltrates are found in all transgenic mouse lines in which behavioral traits are present. Recordings of auditory-evoked potentials in mice of one of these lines demonstrate that transgenic mice are deaf; in these mice spiral ganglia degenerate and most of the cochlear hair cells are absent. By using an S1 nuclease protection assay, we have detected RNA expression of the transgene in all tissues examined and, in particular, at high levels in brain. In situ hybridization experiments show that Mos expression can be detected in specific areas of the central nervous system. Lesions are present in areas with demonstrable overexpression of Mos.

  6. Amount of mobilizable stem cells in perturbed hemopoiesis

    SciTech Connect

    Gidali, J.; Feher, I.

    1985-05-01

    The level of mobilizable 9-day colony-forming units (CFU-s), which represents a constant fraction of the normal mouse bone marrow CFU-s pool, was assayed in BDF1 mice with perturbed hemopoiesis (i.e., during increased turnover of CFU-s or increased CFU-s traffic after irradiation). After low-level irradiation, regeneration of the mobilizable CFU-s fraction was significantly slower than that of bone marrow CFU-s. Depletion of the mobilizable CFU-s pool was observed if a permanently increased outflow of CFU-s from the bone marrow was induced by endotoxin injection. After 40% withdrawal of the blood volume, the mobilizable CFU-s pool expanded marginally. Assuming that the level of mobilizable CFU-s is a consequence of production and outflow from the bone marrow compartment, changes in the pool size of mobilizable CFU-s may be a sensitive indicator of balanced or unbalanced hemopoiesis.

  7. Transgenic lettuce seedlings carrying hepatitis B virus antigen HBsAg.

    PubMed

    Marcondes, Jackson; Hansen, Ekkehard

    2008-12-01

    The obtainment of transgenic edible plants carrying recombinant antigens is a desired issue in search for economic alternatives viewing vaccine production. Here we report a strategy for genetic transformation of lettuce plants (Lactuca sativa L.) using the surface antigen HBsAg of hepatitis B virus. Transgenic lettuce seedlings were obtained through the application of a regulated balance of plant growth regulators. Genetic transformation process was acquired by cocultivation of cotyledons with Agrobacterium tumefaciens harboring the recombinant plasmid. It is the first description of a lettuce Brazilian variety 'Vitória de Verão' genetically modified.

  8. A Mutation of the Prdm9 Mouse Hybrid Sterility Gene Carried by a Transgene.

    PubMed

    Mihola, O; Trachtulec, Z

    2017-01-01

    PRDM9 is a protein with histone-3-methyltransferase activity, which specifies the sites of meiotic recombination in mammals. Deficiency of the Prdm9 gene in the laboratory mouse results in complete arrest of the meiotic prophase of both sexes. Moreover, the combination of certain PRDM9 alleles from different mouse subspecies causes hybrid sterility, e.g., the male-specific meiotic arrest found in the (PWD/Ph × C57BL/6J)F1 animals. The fertility of all these mice can be rescued using a Prdm9-containing transgene. Here we characterized a transgene made from the clone RP24-346I22 that was expected to encompass the entire Prdm9 gene. Both (PWD/Ph × C57BL/6J)F1 intersubspecific hybrid males and Prdm9-deficient laboratory mice of both sexes carrying this transgene remained sterile, suggesting that Prdm9 inactivation occurred in the Tg(RP24-346I22) transgenics. Indeed, comparative qRT-PCR analysis of testicular RNAs from transgene-positive versus negative animals revealed similar expression levels of Prdm9 mRNAs from the exons encoding the C-terminal part of the protein but elevated expression from the regions coding for the N-terminus of PRDM9, indicating that the transgenic carries a new null Prdm9 allele. Two naturally occurring alternative Prdm9 mRNA isoforms were overexpressed in Tg(RP24-346I22), one formed via splicing to a 3'-terminal exon consisting of short interspersed element B2 and one isoform including an alternative internal exon of 28 base pairs. However, the overexpression of these alternative transcripts was apparently insufficient for Prdm9 function or for increasing the fertility of the hybrid males.

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

    PubMed

    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.

  10. Fitness of transgenic mosquito Aedes aegypti males carrying a dominant lethal genetic system.

    PubMed

    Massonnet-Bruneel, Blandine; Corre-Catelin, Nicole; Lacroix, Renaud; Lees, Rosemary S; Hoang, Kim Phuc; Nimmo, Derric; Alphey, Luke; Reiter, Paul

    2013-01-01

    OX513A is a transgenic strain of Aedes aegypti engineered to carry a dominant, non-sex-specific, late-acting lethal genetic system that is repressed in the presence of tetracycline. It was designed for use in a sterile-insect (SIT) pest control system called RIDL® (Release of Insects carrying a Dominant Lethal gene) by which transgenic males are released in the field to mate with wild females; in the absence of tetracycline, the progeny from such matings will not survive. We investigated the mating fitness of OX513A in the laboratory. Male OX513A were as effective as Rockefeller (ROCK) males at inducing refractoriness to further mating in wild type females and there was no reduction in their ability to inseminate multiple females. They had a lower mating success but yielded more progeny than the wild-type comparator strain (ROCK) when one male of each strain was caged with a ROCK female. Mating success and fertility of groups of 10 males-with different ratios of RIDL to ROCK-competing for five ROCK females was similar, but the median longevity of RIDL males was somewhat (18%) lower. We conclude that the fitness under laboratory conditions of OX513A males carrying a tetracycline repressible lethal gene is comparable to that of males of the wild-type comparator strain.

  11. Influence of Nitrogen Availability on Growth of Two Transgenic Birch Species Carrying the Pine GS1a Gene.

    PubMed

    Lebedev, Vadim G; Kovalenko, Nina P; Shestibratov, Konstantin A

    2017-01-06

    An alternative way to increase plant productivity through the use of nitrogen fertilizers is to improve the efficiency of nitrogen utilization via genetic engineering. The effects of overexpression of pine glutamine synthetase (GS) gene and nitrogen availability on growth and leaf pigment levels of two Betula species were studied. Untransformed and transgenic plants of downy birch (B. pubescens) and silver birch (B. pendula) were grown under open-air conditions at three nitrogen regimes (0, 1, or 10 mM) for one growing season. The transfer of the GS1a gene led to a significant increase in the height of only two transgenic lines of nine B. pubescens, but three of five B. pendula transgenic lines were higher than the controls. In general, nitrogen supply reduced the positive effect of the GS gene on the growth of transgenic birch plants. No differences in leaf pigment levels between control and transgenic plants were found. Nitrogen fertilization increased leaf chlorophyll content in untransformed plants but its effect on most of the transgenic lines was insignificant. The results suggest that birch plants carrying the GS gene use nitrogen more efficiently, especially when growing in nitrogen deficient soil. Transgenic lines were less responsive to nitrogen supply in comparison to wild-type plants.

  12. Influence of Nitrogen Availability on Growth of Two Transgenic Birch Species Carrying the Pine GS1a Gene

    PubMed Central

    Lebedev, Vadim G.; Kovalenko, Nina P.; Shestibratov, Konstantin A.

    2017-01-01

    An alternative way to increase plant productivity through the use of nitrogen fertilizers is to improve the efficiency of nitrogen utilization via genetic engineering. The effects of overexpression of pine glutamine synthetase (GS) gene and nitrogen availability on growth and leaf pigment levels of two Betula species were studied. Untransformed and transgenic plants of downy birch (B. pubescens) and silver birch (B. pendula) were grown under open-air conditions at three nitrogen regimes (0, 1, or 10 mM) for one growing season. The transfer of the GS1a gene led to a significant increase in the height of only two transgenic lines of nine B. pubescens, but three of five B. pendula transgenic lines were higher than the controls. In general, nitrogen supply reduced the positive effect of the GS gene on the growth of transgenic birch plants. No differences in leaf pigment levels between control and transgenic plants were found. Nitrogen fertilization increased leaf chlorophyll content in untransformed plants but its effect on most of the transgenic lines was insignificant. The results suggest that birch plants carrying the GS gene use nitrogen more efficiently, especially when growing in nitrogen deficient soil. Transgenic lines were less responsive to nitrogen supply in comparison to wild-type plants. PMID:28067821

  13. Transgenic mice carrying the human poliovirus receptor: new animal models for study of poliovirus neurovirulence.

    PubMed Central

    Horie, H; Koike, S; Kurata, T; Sato-Yoshida, Y; Ise, I; Ota, Y; Abe, S; Hioki, K; Kato, H; Taya, C

    1994-01-01

    Recombinant viruses between the virulent Mahoney and attenuated Sabin 1 strains of poliovirus type 1 were subjected to neurovirulence tests using a transgenic (Tg) mouse line, ICR-PVRTg1, that carried the human poliovirus receptor gene. The Tg mice were inoculated intracerebrally with these recombinant viruses and observed for clinical signs, histopathological lesions, and viral antigens as parameters of neurovirulence of the viruses. These parameters observed in the Tg mice were different for different inoculated viruses. Dose-dependent incidences of paralysis and of death were observed in the Tg mice inoculated with any viruses used. This indicates that values of 50% lethal dose are useful to score a wide range of neurovirulence of poliovirus. The neurovirulence of individual viruses estimated by the Tg mouse model had a strong correlation with those estimated by monkey model. Consequently, the mouse tests identified the neurovirulence determinants on the genome of poliovirus that had been identified by monkey tests. In addition, the mouse tests revealed new neurovirulence determinants, that is, different nucleotides between the two strains at positions 189 and 21 and/or 935 in the 5'-proximal 1,122 nucleotides. The Tg mice used in this study may be suitable for replacing monkeys for investigating poliovirus neurovirulence. Images PMID:8289371

  14. High blood pressure in transgenic mice carrying the rat angiotensinogen gene.

    PubMed Central

    Kimura, S; Mullins, J J; Bunnemann, B; Metzger, R; Hilgenfeldt, U; Zimmermann, F; Jacob, H; Fuxe, K; Ganten, D; Kaling, M

    1992-01-01

    Transgenic mice were generated by injecting the entire rat angiotensinogen gene into the germline of NMRI mice. The resulting transgenic animals were characterized with respect to hemodynamics, parameters of the renin angiotension system, and expression of the transgene. The transgenic line TGM(rAOGEN)123 developed hypertension with a mean arterial blood pressure of 158 mmHg in males and 132 mmHg in females. In contrast, the transgenic line TGM(rAOGEN)92 was not hypertensive. Rat angiotensinogen was detectable only in plasma of animals of line 123. Total plasma angiotensinogen and plasma angiotensin II concentrations were about three times as high as those of negative control mice. In TGM(rAOGEN)123 the transgene was highly expressed in liver and brain. Transcripts were also detected in heart, kidney and testis. In TGM(rAOGEN)92 the brain was the main expressing organ. In situ hybridization revealed an mRNA distribution in the brain of TGM(rAOGEN)123 similar to the one in rat. In TGM(rAOGEN)92 the expression pattern in the brain was aberrant. These data indicate that overexpression of the angiotensinogen gene in liver and brain leads to the development of hypertension in transgenic mice. The TGM(rAOGEN)123 constitutes a high angiotensin II type of hypertension and may provide a new experimental animal model to study the kinetics and function of the renin angiotensin system. Images PMID:1547785

  15. Effect of Vandetanib on Lung Tumorigenesis in Transgenic Mice Carrying an Activating Egfr Gene Mutation.

    PubMed

    Osawa, Masahiro; Ohashi, Kadoaki; Kubo, Toshio; Ichihara, Eiki; Takata, Saburo; Takigawa, Nagio; Takata, Minoru; Tanimoto, Mitsune; Kiura, Katsuyuki

    2016-08-01

    Vandetanib (ZactimaTM) is a novel, orally available inhibitor of both vascular endothelial growth factor receptor-2 (VEGFR-2) and epidermal growth factor receptor (EGFR) tyrosine kinase. In the present study, a line of transgenic mice with a mouse Egfr gene mutation (delE748-A752) corresponding to a human EGFR mutation (delE746-A750) was established. The transgenic mice developed atypical adenomatous hyperplasia to adenocarcinoma of the lung at around 5 weeks of age and died of lung tumors at approximately 17 weeks of age. In the mice treated with vandetanib (6mg/kg/day), these lung tumors disappeared and the phosphorylations of EGFR and VEGFR-2 were reduced in lung tissues to levels comparable to those of non-transgenic control mice. The median overall survival time of the transgenic mice was 28 weeks in the vandetanib-treated group and 17 weeks in the vehicle-treated group. Vandetanib significantly prolonged the survival of the transgenic mice (log-rank test, p< 0.01); resistance to vandetanib occurred at 20 weeks of age and the animals died from their lung tumors at about 28 weeks of age. These data suggest that vandetanib could suppress the progression of tumors harboring an activating EGFR mutation.

  16. c-RET Molecule in Malignant Melanoma from Oncogenic RET-Carrying Transgenic Mice and Human Cell Lines

    PubMed Central

    Takeda, Kozue; Iida, Machiko; Kumasaka, Mayuko; Matsumoto, Yoshinari; Kato, Masashi

    2010-01-01

    Malignant melanoma is one of the most aggressive cancers and its incidence worldwide has been increasing at a greater rate than that of any other cancer. We previously reported that constitutively activated RFP-RET-carrying transgenic mice (RET-mice) spontaneously develop malignant melanoma. In this study, we showed that expression levels of intrinsic c-Ret, glial cell line-derived neurotrophic factor (Gdnf) and Gdnf receptor alpha 1 (Gfra1) transcripts in malignant melanomas from RET-transgenic mice were significantly upregulated compared with those in benign melanocytic tumors. These results suggest that not only introduced oncogenic RET but also intrinsic c-Ret/Gdnf are involved in murine melanomagenesis in RET-mice. We then showed that c-RET and GDNF transcript expression levels in human malignant melanoma cell lines (HM3KO and MNT-1) were higher than those in primary cultured normal human epithelial melanocytes (NHEM), while GFRa1 transcript expression levels were comparable among NHEM, HM3KO and MNT-1. We next showed c-RET and GFRa1 protein expression in HM3KO cells and GDNF-mediated increased levels of their phosphorylated c-RET tyrosine kinase and signal transduction molecules (ERK and AKT) sited potentially downstream of c-RET. Taken together with the finding of augmented proliferation of HM3KO cells after GDNF stimulation, our results suggest that GDNF-mediated c-RET kinase activation is associated with the pathogenesis of malignant melanoma. PMID:20422010

  17. Limbic epilepsy in transgenic mice carrying a Ca2+/calmodulin-dependent kinase II alpha-subunit mutation.

    PubMed Central

    Butler, L S; Silva, A J; Abeliovich, A; Watanabe, Y; Tonegawa, S; McNamara, J O

    1995-01-01

    Multifunctional Ca2+/calmodulin-dependent protein kinase II (CaMK) phosphorylates proteins pivotally involved in diverse neuronal processes and thereby coordinates cellular responses to external stimuli that regulate intracellular Ca2+ [Hanson, P. I. & Schulman, H. (1992) Annu. Rev. Biochem. 61, 559-664]. Despite extensive study, the impact of this enzyme on control of the excitability of neuron populations in the mammalian nervous system in situ is unknown. To address this question, we studied transgenic mice carrying a null mutation (-/-) for the alpha subunit of CaMK. In contrast to wild-type littermates, null mutants exhibit profound hyperexcitability, evident in epileptic seizures involving limbic structures including the hippocampus. No evidence of increased excitability was detected in mice carrying null mutations of the gamma isoform of protein kinase C, underscoring the specificity of the effect of CaMK. CaMK plays a powerful and previously underappreciated role in control of neuronal excitability in the mammalian nervous system. These insights have important implications for analyses of mechanisms of epilepsy and, perhaps, learning and memory. Images Fig. 2 PMID:7624331

  18. An edible vaccine for malaria using transgenic tomatoes of varying sizes, shapes and colors to carry different antigens.

    PubMed

    Chowdhury, Kamal; Bagasra, Omar

    2007-01-01

    different antigens as a single dose. Besides, if immunization schedules could be arranged, the stability of vaccines carrying different malarial antigens, their transport, and the logistics of vaccination would be an almost impossible task to achieve under the current fiscal constraints. We are proposing a unique way to circumvent these logistical difficulties to deliver the malaria vaccines to every susceptible home at a small fraction of a cost. We hypothesize that the anti-malaria edible vaccines in transgenic tomato plants where different transgenic plants expressing different antigenic type(s). Immunizing individuals against 2-3 antigens and against each stage of the life cycle of the multistage parasites would be an efficient, inexpensive and safe way of vaccination. Tomatoes with varying sizes, shapes and colors carrying different antigens would make the vaccines easily identifiable by lay individuals.

  19. A Glimpse into the World of Integrative and Mobilizable Elements in Streptococci Reveals an Unexpected Diversity and Novel Families of Mobilization Proteins.

    PubMed

    Coluzzi, Charles; Guédon, Gérard; Devignes, Marie-Dominique; Ambroset, Chloé; Loux, Valentin; Lacroix, Thomas; Payot, Sophie; Leblond-Bourget, Nathalie

    2017-01-01

    Recent analyses of bacterial genomes have shown that integrated elements that transfer by conjugation play an essential role in horizontal gene transfer. Among these elements, the integrative and mobilizable elements (IMEs) are known to encode their own excision and integration machinery, and to carry all the sequences or genes necessary to hijack the mating pore of a conjugative element for their own transfer. However, knowledge of their prevalence and diversity is still severely lacking. In this work, an extensive analysis of 124 genomes from 27 species of Streptococcus reveals 144 IMEs. These IMEs encode either tyrosine or serine integrases. The identification of IME boundaries shows that 141 are specifically integrated in 17 target sites. The IME-encoded relaxases belong to nine superfamilies, among which four are previously unknown in any mobilizable or conjugative element. A total of 118 IMEs are found to encode a non-canonical relaxase related to rolling circle replication initiators (belonging to the four novel families or to MobT). Surprisingly, among these, 83 encode a TcpA protein (i.e., a non-canonical coupling protein (CP) that is more closely related to FtsK than VirD4) that was not previously known to be encoded by mobilizable elements. Phylogenetic analyses reveal not only many integration/excision module replacements but also losses, acquisitions or replacements of TcpA genes between IMEs. This glimpse into the still poorly known world of IMEs reveals that mobilizable elements have a very high prevalence. Their diversity is even greater than expected, with most encoding a CP and/or a non-canonical relaxase.

  20. A Glimpse into the World of Integrative and Mobilizable Elements in Streptococci Reveals an Unexpected Diversity and Novel Families of Mobilization Proteins

    PubMed Central

    Coluzzi, Charles; Guédon, Gérard; Devignes, Marie-Dominique; Ambroset, Chloé; Loux, Valentin; Lacroix, Thomas; Payot, Sophie; Leblond-Bourget, Nathalie

    2017-01-01

    Recent analyses of bacterial genomes have shown that integrated elements that transfer by conjugation play an essential role in horizontal gene transfer. Among these elements, the integrative and mobilizable elements (IMEs) are known to encode their own excision and integration machinery, and to carry all the sequences or genes necessary to hijack the mating pore of a conjugative element for their own transfer. However, knowledge of their prevalence and diversity is still severely lacking. In this work, an extensive analysis of 124 genomes from 27 species of Streptococcus reveals 144 IMEs. These IMEs encode either tyrosine or serine integrases. The identification of IME boundaries shows that 141 are specifically integrated in 17 target sites. The IME-encoded relaxases belong to nine superfamilies, among which four are previously unknown in any mobilizable or conjugative element. A total of 118 IMEs are found to encode a non-canonical relaxase related to rolling circle replication initiators (belonging to the four novel families or to MobT). Surprisingly, among these, 83 encode a TcpA protein (i.e., a non-canonical coupling protein (CP) that is more closely related to FtsK than VirD4) that was not previously known to be encoded by mobilizable elements. Phylogenetic analyses reveal not only many integration/excision module replacements but also losses, acquisitions or replacements of TcpA genes between IMEs. This glimpse into the still poorly known world of IMEs reveals that mobilizable elements have a very high prevalence. Their diversity is even greater than expected, with most encoding a CP and/or a non-canonical relaxase. PMID:28373865

  1. Transgenic cattle produced by nuclear transfer of fetal fibroblasts carrying Ipr1 gene at a specific locus.

    PubMed

    Wang, Yong Sheng; He, Xiaoning; Du, Yue; Su, Jianmin; Gao, Mingqing; Ma, Yefei; Hua, Song; Quan, Fusheng; Liu, Jun; Zhang, Yong

    2015-09-01

    This study aimed to assess the effects of the intracellular pathogen resistance 1 (Ipr1) transgene on preventing infection of Mycobacterium bovis in cattle. A specific expression vector for the Ipr1 gene was constructed and inserted in the genome between surfactant protein A and methionine adenosyltransferase I of bovine fetal fibroblasts. After SCNT, cleavage (86.9% vs. 87.4%, P > 0.05) and blastocyst developmental rates (34.6% vs. 33.5%, P > 0.05) were similar between transgenic and nontransgenic bovine fetal fibroblasts. Four surviving and one dead Ipr1-transgenic female cattle were produced by transfer of the SCNT blastocysts. Polymerase chain reaction and Southern blot analyses confirmed that the Ipr1 transgene of the cattle was located at the expected site. Inserting Ipr1 gene did not affect the expression of the surrounding genes. Main death modality of M bovis-infected peripheral blood mononuclear cells (PBMCs) derived from Ipr1-transgenic cattle was apoptosis, whereas that of PBMCs from control cattle was necrosis. In addition, the number of colony-forming units in PBMCs of Ipr1-transgenic cattle was significantly lower than that of the control cattle (P < 0.05). The finding that expression of Ipr1 transgene in PBMCs significantly increased anti-M bovis activity suggested breeding anti-M bovis cattle population by the transgenic SCNT technique could be a feasible strategy.

  2. Bacteroides mobilizable and conjugative genetic elements: antibiotic resistance among clinical isolates.

    PubMed

    Quesada-Gómez, Carlos

    2011-12-01

    The conjugation is one of the most important mechanisms of horizontal gene transfer in prokaryotes, leading to genetic variation within a species and the acquisition of new traits, such as antibiotic resistance. Bacteroides is an obligate anaerobe of the colon and a significant opportunistic pathogen. Antibiotic resistance among Bacteroides spp. is rapidly increasing, largely due to the dissemination of DNA transfer factors (plasmids and transposons) harbored by members of this genus. Transfer factors can be divided into two classes, conjugative and mobilizable. Species of the intestinal Bacteroides have yielded different resistance plasmids, all of which have been intensely studied, the plasmids encode high-level MLS resistance conferred by a conserved erm gene. It has been reported an interesting observation associated with the transfer of several of these types of elements, all of which conferred Tcr and displayed greatly increased transfer efficiency following exposure to tetracycline. Many of the conjugative transposons (CTns) in Bacteroides are related to various genetic elements (such as CTnDOT, CTnERL, NBU and others). CTnDOT carries a tetracycline resistance gene, tetQ, and an erythromycin resistance gene, ermF. Resistance to drugs used to treat Bacteroides infections, such as clindamycin, has also been increasing. These conjugal elements have been found in Bacteroides clinical isolates. Thus, horizontal gene transfer could conceivably have played a role in the rising incidence of resistance in this bacterial group.

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

    PubMed Central

    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 muα-globin2/huβ-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

  4. Spread and exchange of bla NDM-1 in hospitalized neonates: role of mobilizable genetic elements.

    PubMed

    Datta, S; Mitra, S; Chattopadhyay, P; Som, T; Mukherjee, S; Basu, S

    2017-02-01

    To investigate the mobilizable elements associated with bla NDM-1 in Enterobacteriaceae isolated from septicaemic neonates at a NICU in India, during December, 2008-2011. An attempt was also made to understand whether there was a pattern in the temporal acquisition of bla NDM-1 within the unit. Transferability of carbapenem resistance was tested by conjugation and transformation. Plasmid types and addiction systems were analysed. The genetic background of bla NDM-1 and association with class 1 integron were evaluated by PCR mapping. RFLP was carried out to discriminate plasmids of same incompatibility group. Transfer of carbapenem resistance was successful in 13/15 cases. bla NDM-1 was associated with different plasmid scaffolds (IncFII, IncL/M, IncN, IncR, IncHIB-M/FIB-M), IncF type being the prevalent one. Addiction systems ccdAB and hok/sok were associated with transferable plasmids. Genetic structures surrounding bla NDM-1 showed its association with at least a remnant of ISAba125 at its 5'-end. The spread of NDM-1 was not related to class 1 integron which possessed resistance determinants against trimethoprim (dfrA12, dfrA1, dfrA5), streptomycin (aadA2, aacA4), and rifampicin (arr-3). RFLP showed that three isolates possessed the same FII/FIIs plasmid; two of these three isolates were from a single neonate, implying interspecies transfer of bla NDM-1. The predominance of FII plasmids and ISAba125 along with bla NDM-1 was noted, but no specific pattern in the temporal acquisition of mobile genetic elements could be identified. To the best of our knowledge, this report is the first to inform the in-vivo interspecies plasmid transfer event of bla NDM-1 in a neonate.

  5. The mesenchymal stem cells derived from transgenic mice carrying human coagulation factor VIII can correct phenotype in hemophilia A mice.

    PubMed

    Wang, Qing; Gong, Xiuli; Gong, Zhijuan; Ren, Xiaoyie; Ren, Zhaorui; Huang, Shuzhen; Zeng, Yitao

    2013-12-20

    Hemophilia A (HA) is an inherited X-linked recessive bleeding disorder caused by coagulant factor VIII (FVIII) deficiency. Previous studies showed that introduction of mesenchymal stem cells (MSCs) modified by FVIII-expressing retrovirus may result in phenotypic correction of HA animals. This study aimed at the investigation of an alternative gene therapy strategy that may lead to sustained FVIII transgene expression in HA mice. B-domain-deleted human FVIII (hFVIIIBD) vector was microinjected into single-cell embryos of wild-type mice to generate a transgenic mouse line, from which hFVIIIBD-MSCs were isolated, followed by transplantation into HA mice. RT-PCR and real-time PCR analysis demonstrated the expression of hFVIIIBD in multi-organs of recipient HA mice. Immunohistochemistry showed the presence of hFVIIIBD positive staining in multi-organs of recipient HA mice. ELISA indicated that plasma hFVIIIBD level in recipient mice reached its peak (77 ng/mL) at the 3rd week after implantation, and achieved sustained expression during the 5-week observation period. Plasma FVIII activities of recipient HA mice increased from 0% to 32% after hFVIIIBD-MSCs transplantation. APTT (activated partial thromboplastin time) value decreased in hFVIIIBD-MSCs transplanted HA mice compared with untreated HA mice (45.5 s vs. 91.3 s). Our study demonstrated an effective phenotypic correction in HA mice using genetically modified MSCs from hFVIIIBD transgenic mice.

  6. Spontaneous destructive periodontitis and skeletal bone damage in transgenic mice carrying a human shared epitope-coding HLA-DRB1 allele

    PubMed Central

    Gehlot, Prashasnika; Volk, Sarah L; Rios, Hector F; Jepsen, Karl J; Holoshitz, Joseph

    2016-01-01

    Objective Shared epitope (SE)-coding DRB1 alleles are associated with bone erosion in several diseases, including rheumatoid arthritis (RA) and periodontal disease (PD), but the underlying mechanism is unknown. We have recently identified the SE as an osteoclast-activating ligand. To better understand the biological effects of the SE in vivo, here we sought to determine whether it can facilitate spontaneous bone damage in naïve mice. Methods 3-month old naïve transgenic mice that carry the human SE-coding allele DRB1*04:01, or a SE-negative allele DRB1*04:02 were studied. Bone tissues were analysed by micro-CT, and the tooth-supporting tissues were studied by histology, immunohistochemistry and immunofluorescence. Serum biomarkers were determined by ELISA. Results Transgenic mice expressing the SE-coding DRB1*04:01 allele, but not mice carrying the SE-negative allele DRB1*04:02, showed spontaneous PD associated with interleukin (IL)-17 overabundance and periostin disruption. Mandibular bone volumetric and mineralisation parameters were significantly lower in SE-positive mice, and alveolar bone resorption was significantly increased in these mice. SE-positive mice also had more slender tibiae, and their marrow, cortical and total areas were lower than those of SE-negative mice. Additionally, significantly increased serum IL-17, tumour necrosis factor-α and osteoprotegrin levels were found in SE-positive mice, while their receptor activator of nuclear factor κ-B ligand levels were significantly lower. Conclusions A human SE-coding allele increases the propensity to spontaneous bone-destructive periodontal inflammation and skeletal bone damage in transgenic mice. These findings provide new insights into the previously documented but poorly understood association of the SE with accelerated bone erosion in RA and several other human diseases. PMID:27933212

  7. Overexpression of transforming growth factor-. beta. in transgenic mice carrying the human T-cell lymphotropic virus type I tax gene

    SciTech Connect

    Kim, Seongjin; Winokur, T.S.; Lee, Hyde; Danielpour, D.; Kim, Kyung Young; Geiser, A.G.; Sporn, M.B.; Roberts, A.B. ); Chen, Liansheng; Jay, G. )

    1991-10-01

    Human T-cell lymphotropic virus type I (HTLV-I) has been associated with an adult form of T-cell leukemia as well as tropical spastic paraparesis, a neurodegenerative disease. Adult T-cell leukemia patients express high levels of the type 1 isoform of transforming growth factor-{beta} (TGF-{beta}1), which is mediated by the effects of the HTLV-I Tax transactivator protein on the TGF-{beta}1 promoter. To understand further the regulation of TGF-{beta}1 expression by Tax, the authors examined its expression in transgenic mice carrying the HTLV-I tax gene. They show that tumors from these mice and other tissues, such as submaxillary glands and skeletal muscle, which express high levels of tax mRNA selectively express high levels of TGF-{beta}1 mRNA and protein. Moreover, TGF-{beta}1 significantly stimulated the incorporation of tritiated thymidine into one of three cells lines derived from neurofibromas of tax-transgenic mice, which suggest that the excessive production of TGF-{beta}1 may play a role in tumorigenesis and that these mice may serve as a useful model for studying the biological effects of TGF-{beta} in vivo.

  8. Systemic administration of attenuated Salmonella choleraesuis carrying thrombospondin-1 gene leads to tumor-specific transgene expression, delayed tumor growth and prolonged survival in the murine melanoma model.

    PubMed

    Lee, Che-Hsin; Wu, Chao-Liang; Shiau, Ai-Li

    2005-02-01

    Some anaerobic and facultative anaerobic bacteria have been used experimentally as anticancer agents because of their selective growth in the hypoxia regions of solid tumors after systemic administration. We have previously shown the feasibility of using attenuated Salmonella choleraesuis as a gene delivery vector. In this study, we exploited S. choleraesuis carrying thrombospondin-1 (TSP-1) gene for treating primary melanoma and experimental pulmonary metastasis in the syngeneic murine B16F10 melanoma model. Systemic administration of S. choleraesuis allowed targeted gene delivery to tumors. The bacteria accumulated preferentially in tumors over livers and spleens at ratios ranging from 1000:1 to 10,000:1. The level of transgene expression via S. choleraesuis-mediated gene transfer in tumors could reach more than 1800-fold higher than in livers and spleens. Notably, bacterial accumulation was also observed in the lungs with metastatic nodules, but not in healthy lungs. When administered into mice bearing subcutaneous or pulmonary metastatic melanomas, S. choleraesuis carrying TSP-1 gene significantly inhibited tumor growth and enhanced survival of the mice. Immunohistochemical studies in the tumors from these mice displayed decreased intratumoral microvessel density. Taken together, these findings suggest that TSP-1 gene therapy delivered by S. choleraesuis may be effective for the treatment of primary as well as metastatic melanomas.

  9. Analysis of the physiological mechanism of salt-tolerant transgenic rice carrying a vacuolar Na+/H + antiporter gene from Suaeda salsa.

    PubMed

    Zhao, Fengyun; Wang, Zenglan; Zhang, Quan; Zhao, Yanxiu; Zhang, Hui

    2006-03-01

    Salt stress is one of the most serious factors limiting the productivity of agricultural crops. Increasing evidence has demonstrated that vacuolar Na+/H+ antiporters play a crucial role in plant salt tolerance. In the present study, we expressed the Suaeda salsa vacuolar Na+/H+ antiporter SsNHX1 in transgenic rice to investigate whether this can increase the salt tolerance of rice, and to study how overexpression of this gene affected other salt-tolerant mechanisms. It was found that transgenic rice plants showed markedly enhanced tolerance to salt stress and to water deprivation compared with non-transgenic controls upon salt stress imposition under outdoor conditions. Measurements of ion levels indicated that K+, Ca2+ and Mg2+ contents were all higher in transgenic plants than in non-transformed controls. Furthermore, shoot V-ATPase hydrolytic activity was dramatically increased in transgenics compared to that of non-transformed controls under salt stress conditions. Physiological analysis also showed that the photosynthetic activity of the transformed plants was higher whereas the same plants had reduced reactive oxygen species generation. In addition, the soluble sugar content increased in the transgenics compared with that in non-transgenics. These results imply that up-regulation of a vacuolar Na+/H+ antiporter gene in transgenic rice might cause pleiotropic up-regulation of other salt-resistance-related mechanisms to improve salt tolerance.

  10. Mobilizable Rolling-Circle Replicating Plasmids from Gram-Positive Bacteria: A Low-Cost Conjugative Transfer.

    PubMed

    Fernández-López, Cris; Bravo, Alicia; Ruiz-Cruz, Sofía; Solano-Collado, Virtu; Garsin, Danielle A; Lorenzo-Díaz, Fabián; Espinosa, Manuel

    2014-10-01

    Conjugation is a key mechanism for horizontal gene transfer in bacteria. Some plasmids are not self-transmissible but can be mobilized by functions encoded in trans provided by other auxiliary conjugative elements. Although the transfer efficiency of mobilizable plasmids is usually lower than that of conjugative elements, mobilizable plasmids are more frequently found in nature. In this sense, replication and mobilization can be considered important mechanisms influencing plasmid promiscuity. Here we review the currently available information on two families of small mobilizable plasmids from Gram-positive bacteria that replicate via the rolling-circle mechanism. One of these families, represented by the streptococcal plasmid pMV158, is an interesting model since it contains a specific mobilization module (MOBV) that is widely distributed among mobilizable plasmids. We discuss a mechanism in which the promiscuity of the pMV158 replicon is based on the presence of two origins of lagging strand synthesis. The current strategies to assess plasmid transfer efficiency as well as to inhibit conjugative plasmid transfer are presented. Some applications of these plasmids as biotechnological tools are also reviewed.

  11. Mobilizable Rolling-Circle Replicating Plasmids from Gram-Positive Bacteria: A Low-Cost Conjugative Transfer

    PubMed Central

    Fernández-López, Cris; Bravo, Alicia; Ruiz-Cruz, Sofía; Solano-Collado, Virtu; Garsin, Danielle A.; Lorenzo-Díaz, Fabián; Espinosa, Manuel

    2014-01-01

    Chapter summary Conjugation is a key mechanism for horizontal gene transfer in bacteria. Some plasmids are not self-transmissible but can be mobilized by functions encoded in trans provided by other auxiliary conjugative elements. Although the transfer efficiency of mobilizable plasmids is usually lower than that of conjugative elements, mobilizable plasmidsare more frequently found in nature. In this sense, replication and mobilization can be considered as important mechanisms influencing plasmid promiscuity. Here we review the present available information on two families of small mobilizable plasmids from Gram-positive bacteria that replicate via the rolling-circle mechanism. One of these families, represented by the streptococcal plasmid pMV158, is an interesting model since it contains a specific mobilization module (MOBV) that is widely distributed among mobilizable plasmids. We discuss a mechanism in which the promiscuity of the pMV158 replicon is based on the presence of two origins of lagging strand synthesis. The current strategies to assess plasmid transfer efficiency as well as to inhibit conjugative plasmid transfer are presented. Some applications of these plasmids as biotechnological tools are also reviewed. PMID:25606350

  12. Meclozine promotes longitudinal skeletal growth in transgenic mice with achondroplasia carrying a gain-of-function mutation in the FGFR3 gene.

    PubMed

    Matsushita, Masaki; Hasegawa, Satoru; Kitoh, Hiroshi; Mori, Kensaku; Ohkawara, Bisei; Yasoda, Akihiro; Masuda, Akio; Ishiguro, Naoki; Ohno, Kinji

    2015-02-01

    Achondroplasia (ACH) is one of the most common skeletal dysplasias causing short stature owing to a gain-of-function mutation in the FGFR3 gene, which encodes the fibroblast growth factor receptor 3. We found that meclozine, an over-the-counter drug for motion sickness, inhibited elevated FGFR3 signaling in chondrocytic cells. To examine the feasibility of meclozine administration in clinical settings, we investigated the effects of meclozine on ACH model mice carrying the heterozygous Fgfr3(ach) transgene. We quantified the effect of meclozine in bone explant cultures employing limb rudiments isolated from developing embryonic tibiae from Fgfr3(ach) mice. We found that meclozine significantly increased the full-length and cartilaginous primordia of embryonic tibiae isolated from Fgfr3(ach) mice. We next analyzed the skeletal phenotypes of growing Fgfr3(ach) mice and wild-type mice with or without meclozine treatment. In Fgfr3(ach) mice, meclozine significantly increased the body length after 2 weeks of administration. At skeletal maturity, the bone lengths including the cranium, radius, ulna, femur, tibia, and vertebrae were significantly longer in meclozine-treated Fgfr3(ach) mice than in untreated Fgfr3(ach) mice. Interestingly, meclozine also increased bone growth in wild-type mice. The plasma concentration of meclozine during treatment was within the range that has been used in clinical settings for motion sickness. Increased longitudinal bone growth in Fgfr3(ach) mice by oral administration of meclozine in a growth period suggests potential clinical feasibility of meclozine for the improvement of short stature in ACH.

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

    PubMed

    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.

  14. Ectopic bone formation and chondrodysplasia in transgenic mice carrying the rat C3(1)/T{sub AG} fusion gene

    SciTech Connect

    Green, J.E.; Maroulakou, I.G.; Anver, M.

    1994-09-01

    Transgenic mice expressing the SV40 large T-antigen (T{sup AG}) under the regultory control of the hormone-responsive rat C3(1) prostatein promoter develop unusual bone and cartilage lesions, as well as ectopic bone and cartilage formation. Two lines of transgenic animals have been propagated in which the expression of the transgene in chondrocytes results in a mild to moderate generalized disorganization of cartilage growth which appears to affect multiple tissues, including the trachea, ear pinna and articular cartilage. The epiphyseal plates are also affected with normal architecture of the zones of proliferation and maturation, but marked elongation of the zone of hypertrophy. Immunocytochemistry demonstrates that expression of T{sup AG} is limited to the zone of hypertropny in the epiphyseal plates, suggesting that the chondrocytes become hormone-responsive at this particular stage of differentiation. Normal mineralization and trabecular formation in long bone appears to occur. Ectopic bone and cartilage formation occurs in the foot pads of the fore- and hind- feet over the course of several months. This is preceded by proliferation of sweat gland epithelial cells followed by the appearance of nodules of cartilage and bone. The nodules are closely associated with proliferating epithelium but are not contiguous with bony structures normally found in the feet. The roles of BMP`s, growth factors, oncogenes and hormones in the development of these lesions will be presented. These transgenic animals may provide new insights into hormone-responsiveness of chondrocytes, as well as factors involved in the processes of bone and cartilage differentiation and growth. These transgenic animals may serve as a useful model for human heterotopic bone formation.

  15. Streptococcal group B integrative and mobilizable element IMESag-rpsI encodes a functional relaxase involved in its transfer

    PubMed Central

    Lorenzo-Diaz, Fabian; Fernández-Lopez, Cris; Douarre, Pierre-Emmanuel; Baez-Ortega, Adrian; Flores, Carlos; Glaser, Philippe

    2016-01-01

    Streptococcus agalactiae or Group B Streptococcus (GBS) are opportunistic bacteria that can cause lethal sepsis in children and immuno-compromised patients. Their genome is a reservoir of mobile genetic elements that can be horizontally transferred. Among them, integrative and conjugative elements (ICEs) and the smaller integrative and mobilizable elements (IMEs) primarily reside in the bacterial chromosome, yet have the ability to be transferred between cells by conjugation. ICEs and IMEs are therefore a source of genetic variability that participates in the spread of antibiotic resistance. Although IMEs seem to be the most prevalent class of elements transferable by conjugation, they are poorly known. Here, we have studied a GBS-IME, termed IMESag-rpsI, which is widely distributed in GBS despite not carrying any apparent virulence trait. Analyses of 240 whole genomes showed that IMESag-rpsI is present in approximately 47% of the genomes, has a roughly constant size (approx. 9 kb) and is always integrated at a single location, the 3′-end of the gene encoding the ribosomal protein S9 (rpsI). Based on their genetic variation, several IMESag-rpsI types were defined (A–J) and classified in clonal complexes (CCs). CC1 was the most populated by IMESag-rpsI (more than 95%), mostly of type-A (71%). One CC1 strain (S. agalactiae HRC) was deep-sequenced to understand the rationale underlying type-A IMESag-rpsI enrichment in GBS. Thirteen open reading frames were identified, one of them encoding a protein (MobSag) belonging to the broadly distributed family of relaxases MOBV1. Protein MobSag was purified and, by a newly developed method, shown to cleave DNA at a specific dinucleotide. The S. agalactiae HRC-IMESag-rpsI is able to excise from the chromosome, as shown by the presence of circular intermediates, and it harbours a fully functional mobilization module. Further, the mobSag gene encoded by this mobile element is able to promote plasmid transfer among pneumococcal

  16. Expression Analysis of Hairpin RNA Carrying Sugarcane mosaic virus (SCMV) Derived Sequences and Transgenic Resistance Development in a Model Rice Plant

    PubMed Central

    Akbar, Sehrish; Wang, Ming-Bo; Liu, Qing

    2017-01-01

    Developing transgenic resistance in monocotyledonous crops against pathogens remains a challenging area of research. Sugarcane mosaic virus (SCMV) is a serious pathogen of many monocotyledonous crops including sugarcane. The objective of present study was to analyze transgenic expression of hairpin RNA (hpRNA), targeting simultaneously CP (Coat Protein) and Hc-Pro (helper component-proteinase) genes of SCMV, in a model rice plant. Conserved nucleotide sequences, exclusive for DAG (Aspartic acid-Alanine-Glycine) and KITC (Lycine-Isoleucine-Threonine-Cysteine) motifs, derived from SCMV CP and Hc-Pro genes, respectively, were fused together and assembled into the hpRNA cassette under maize ubiquitin promoter to form Ubi-hpCP:Hc-Pro construct. The same CP:Hc-Pro sequence was fused with the β-glucuronidase gene (GUS) at the 3′ end under CaMV 35S promoter to develop 35S-GUS:CP:Hc-Pro served as a target reporter gene construct. When delivered into rice callus tissues by particle bombardment, the Ubi-hpCP:Hc-Pro construct induced strong silencing of 35S-GUS:CP:Hc-Pro. Transgenic rice plants, containing Ubi-hpCP:Hc-Pro construct, expressed high level of 21–24 nt small interfering RNAs, which induced specific suppression against GUS:CP:Hc-Pro delivered by particle bombardment and conferred strong resistance to mechanically inoculated SCMV. It is concluded that fusion hpRNA approach is an affordable method for developing resistance against SCMV in model rice plant and it could confer SCMV resistance when transformed into sugarcane. PMID:28255554

  17. Transfer of mupirocin resistance from Staphylococcus haemolyticus clinical strains to Staphylococcus aureus through conjugative and mobilizable plasmids.

    PubMed

    Rossi, Ciro C; Ferreira, Natália C; Coelho, Marcus L V; Schuenck, Ricardo P; Bastos, Maria do Carmo de F; Giambiagi-deMarval, Marcia

    2016-07-01

    Coagulase-negative staphylococci are thought to act as reservoirs of antibiotic resistance genes that can be transferred to Staphylococcus aureus, thus hindering the combat of this bacterium. In this work, we analyzed the presence of plasmids conferring resistance to the antibiotic mupirocin-widely used to treat and prevent S. aureus infections in hospital environments-in nosocomial S. haemolyticus strains. About 12% of the 75 strains tested were resistant to mupirocin, and this phenotype was correlated with the presence of plasmids. These plasmids were shown to be diverse, being either conjugative or mobilizable, and capable of transferring mupirocin resistance to S. aureus Our findings reinforce that S. haemolyticus, historically and mistakenly considered as a less important pathogen, is a reservoir of resistance genes which can be transferred to other bacteria, such as S. aureus, emphasizing the necessity of more effective strategies to detect and combat this emergent opportunistic pathogen.

  18. Rapid growth of invasive metastatic melanoma in carcinogen-treated hepatocyte growth factor/scatter factor-transgenic mice carrying an oncogenic CDK4 mutation.

    PubMed

    Tormo, Damia; Ferrer, Aleix; Gaffal, Evelyn; Wenzel, Jörg; Basner-Tschakarjan, Etiena; Steitz, Julia; Heukamp, Lukas C; Gütgemann, Ines; Buettner, Reinhard; Malumbres, Marcos; Barbacid, Mariano; Merlino, Glenn; Tüting, Thomas

    2006-08-01

    Currently, novel mouse models of melanoma are being generated that recapitulate the histopathology and molecular pathogenesis observed in human disease. Impaired cell-cycle control, which is a hallmark of both familial and sporadic melanoma, promotes slowly growing carcinogen-induced melanomas in the skin of mice carrying a mutated cyclin-dependent kinase 4 (CDK4(R24C)). Deregulated receptor tyrosine kinase signaling, which is another important feature of human melanoma, leads to spontaneous development of metastatic melanoma after a long latency period in mice overexpressing hepatocyte growth factor/scatter factor (HGF/SF mice). Here we report that treatment with 7,12-dimethylbenz[a]anthracene and 12-O-tetradecanoylphorbol-13-acetate induced metastatic melanomas in all HGF/SF mice on the C57BL/6 background, which histologically resemble human melanoma. Importantly, mutant CDK4 dramatically increased the number and the growth kinetics of carcinogen-induced primary melanomas in the skin and promoted the growth of spontaneous metastases in lymph nodes and lungs in all HGF/SF mice within the first 3 months of life. Apart from very few skin papillomas, we did not observe tumors of other histology in carcinogen-treated HGF/SF x CDK4(R24C) mice. This new experimental mouse model can now be exploited to study further the biology of melanoma and evaluate new treatment modalities.

  19. trans activation of granulocyte-macrophage colony-stimulating factor and the interleukin-2 receptor in transgenic mice carrying the human T-lymphotropic virus type 1 tax gene.

    PubMed

    Green, J E; Begley, C G; Wagner, D K; Waldmann, T A; Jay, G

    1989-11-01

    Three lines of transgenic mice carrying the human T-cell lymphotropic virus type 1 tax gene have previously been reported to develop neurofibromas composed of perineural fibroblasts (S. H. Hinrichs, M. Nerenberg, R. K. Reynolds, G. Khoury, and G. Jay, Science 237:1340-1343, 1987; M. Nerenberg, S. H. Hinrichs, R. K. Reynolds, G. Khoury, and G. Jay, Science 237:1324-1329, 1987). Tumors from these mice and tumor cell lines derived from them expressed high levels of tax RNA and protein. They also expressed high levels of the granulocyte-macrophage colony-stimulating factor (GM-CSF) gene as measured by proliferative responses of FD-CP1 target cells using conditioned media from tumor cells and by Northern (RNA) blot analysis of RNA from tumors and tumor cell lines. Although other tissues, such as salivary glands and muscles, in the transgenic mice also expressed high levels of tax, they did not express the gene for GM-CSF. This indicates that tissue-specific cellular factors, in addition to tax, are required for GM-CSF gene expression. Systemic effects of excessive GM-CSF production were demonstrated by infiltration of polymorphonuclear leukocytes into tumor tissues which are not necrotic, by peripheral granulocytosis, and by splenomegaly resulting from myeloid hyperplasia. The interleukin-2 (IL-2) receptor was also found to be expressed by the tumors and tumor cell lines as measured by IL-2-binding and cross-linking studies. This is the first demonstration that the IL-2 receptor can be activated by tax in a nonlymphoid cell type. These in vivo findings are consistent with other reports which have demonstrated in vitro cis-regulatory elements within the 5'-flanking regions of the genes for GM-CSF and the IL-2 receptor which are responsive to trans activation by the tax gene.

  20. A toxin antitoxin system promotes the maintenance of the IncA/C-mobilizable Salmonella Genomic Island 1.

    PubMed

    Huguet, Kevin T; Gonnet, Mathieu; Doublet, Benoît; Cloeckaert, Axel

    2016-08-31

    The multidrug resistance Salmonella Genomic Island 1 (SGI1) is an integrative mobilizable element identified in several enterobacterial pathogens. This chromosomal island requires a conjugative IncA/C plasmid to be excised as a circular extrachromosomal form and conjugally mobilized in trans. Preliminary observations suggest stable maintenance of SGI1 in the host chromosome but paradoxically also incompatibility between SGI1 and IncA/C plasmids. Here, using a Salmonella enterica serovar Agona clonal bacterial population as model, we demonstrate that a Toxin-Antitoxin (TA) system encoded by SGI1 plays a critical role in its stable host maintenance when an IncA/C plasmid is concomitantly present. This system, designated sgiAT for Salmonella genomic island 1 Antitoxin and Toxin respectively, thus seems to play a stabilizing role in a situation where SGI1 is susceptible to be lost through plasmid IncA/C-mediated excision. Moreover and for the first time, the incompatibility between SGI1 and IncA/C plasmids was experimentally confirmed.

  1. A toxin antitoxin system promotes the maintenance of the IncA/C-mobilizable Salmonella Genomic Island 1

    PubMed Central

    Huguet, Kevin T.; Gonnet, Mathieu; Doublet, Benoît; Cloeckaert, Axel

    2016-01-01

    The multidrug resistance Salmonella Genomic Island 1 (SGI1) is an integrative mobilizable element identified in several enterobacterial pathogens. This chromosomal island requires a conjugative IncA/C plasmid to be excised as a circular extrachromosomal form and conjugally mobilized in trans. Preliminary observations suggest stable maintenance of SGI1 in the host chromosome but paradoxically also incompatibility between SGI1 and IncA/C plasmids. Here, using a Salmonella enterica serovar Agona clonal bacterial population as model, we demonstrate that a Toxin-Antitoxin (TA) system encoded by SGI1 plays a critical role in its stable host maintenance when an IncA/C plasmid is concomitantly present. This system, designated sgiAT for Salmonella genomic island 1 Antitoxin and Toxin respectively, thus seems to play a stabilizing role in a situation where SGI1 is susceptible to be lost through plasmid IncA/C-mediated excision. Moreover and for the first time, the incompatibility between SGI1 and IncA/C plasmids was experimentally confirmed. PMID:27576575

  2. Carrying Loom

    ERIC Educational Resources Information Center

    Lazaron, Edna

    1976-01-01

    Whenever a young student wanted to weave, his loom was at school or at home. He solved the problem by designing a portable loom which he is able to carry with his school books and can even use on the school bus. (Author/RK)

  3. Transgenic Animals.

    ERIC Educational Resources Information Center

    Jaenisch, Rudolf

    1988-01-01

    Describes three methods and their advantages and disadvantages for introducing genes into animals. Discusses the predictability and tissue-specificity of the injected genes. Outlines the applications of transgenic technology for studying gene expression, the early stages of mammalian development, mutations, and the molecular nature of chromosomes.…

  4. Analysis of pMA67, a predicted rolling-circle replicating, mobilizable, tetracycline-resistance plasmid from the honey bee pathogen, Paenibacillus larvae.

    PubMed

    Murray, K Daniel; Aronstein, Katherine A; de León, Jesse H

    2007-09-01

    This work characterizes a recently discovered natural tetracycline-resistance plasmid called pMA67 from Paenibacillus larvae--a Gram-positive bacterial pathogen of honey bees. We provide evidence that pMA67 replicates by the rolling-circle mechanism, and sequence comparisons place it in the pMV158 family of rolling-circle replicons. The plasmid contains predicted rep, cop, and rnaII genes for control of replication initiating at a predicted double-strand origin. The plasmid has an ssoT single-strand origin, which is efficient enough to allow only very small amounts of the single-stranded DNA intermediate to accumulate. The overall efficiency of replication is sufficient to render the plasmid segregationally stable without selection in P. larvae and in Bacillus megaterium, but not in Escherichia coli. The plasmid is expected to be mobilizable due to the presence of a mob gene and an oriT site. The plasmid contains a tetL gene, whose predicted amino acid sequence implies a relatively ancient divergence from all previously known plasmid-encoded tetL genes. We confirm that the tetL gene alone is sufficient for conferring resistance to tetracyclines. Sequence comparisons, mostly with the well-characterized pMV158, allow us to predict promoters, DNA and RNA secondary structures, DNA and protein motifs, and other elements.

  5. Transgenic animal bioreactors.

    PubMed

    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

  6. Transgenic mouse offspring generated by ROSI.

    PubMed

    Moreira, Pedro; Pérez-Cerezales, Serafín; Laguna, Ricardo; Fernández-Gonzalez, Raúl; Sanjuanbenito, Belén Pintado; Gutiérrez-Adán, Alfonso

    2016-01-01

    The production of transgenic animals is an important tool for experimental and applied biology. Over the years, many approaches for the production of transgenic animals have been tried, including pronuclear microinjection, sperm-mediated gene transfer, transfection of male germ cells, somatic cell nuclear transfer and the use of lentiviral vectors. In the present study, we developed a new transgene delivery approach, and we report for the first time the production of transgenic animals by co-injection of DNA and round spermatid nuclei into non-fertilized mouse oocytes (ROSI). The transgene used was a construct containing the human CMV immediate early promoter and the enhanced GFP gene. With this procedure, 12% of the live offspring we obtained carried the transgene. This efficiency of transgenic production by ROSI was similar to the efficiency by pronuclear injection or intracytoplasmic injection of male gamete nuclei (ICSI). However, ICSI required fewer embryos to produce the same number of transgenic animals. The expression of Egfp mRNA and fluorescence of EGFP were found in the majority of the organs examined in 4 transgenic lines generated by ROSI. Tissue morphology and transgene expression were not distinguishable between transgenic animals produced by ROSI or pronuclear injection. Furthermore, our results are of particular interest because they indicate that the transgene incorporation mediated by intracytoplasmic injection of male gamete nuclei is not an exclusive property of mature sperm cell nuclei with compact chromatin but it can be accomplished with immature sperm cell nuclei with decondensed chromatin as well. The present study also provides alternative procedures for transgene delivery into embryos or reconstituted oocytes.

  7. Variegated transgene expression in mouse mammary gland is determined by the transgene integration locus.

    PubMed Central

    Dobie, K W; Lee, M; Fantes, J A; Graham, E; Clark, A J; Springbett, A; Lathe, R; McClenaghan, M

    1996-01-01

    Mice carrying an ovine beta-lactoglobulin (BLG) transgene secrete BLG protein into their milk. To explore transgene expression stability, we studied expression levels in three BLG transgenic mouse lines. Unexpectedly, two lines exhibited variable levels of transgene expression. Copy number within lines appeared to be stable and there was no evidence of transgene rearrangement. In the most variable line, BLG production levels were stable within individual mice in two successive lactations. Backcrossing demonstrated that genetic background did not contribute significantly to variable expression. Tissue in situ hybridization revealed mosaicism of transgene expression within individual mammary glands from the two variable lines; in low expressors, discrete patches of cells expressing the transgene were observed. Transgene protein concentrations in milk reflected the proportion of epithelial cells expressing BLG mRNA. Furthermore, chromosomal in situ hybridization revealed that transgene arrays in both lines are situated close to the centromere. We propose that mosaicism of transgene expression is a consequence of the chromosomal location and/or the nature of the primary transgene integration event. Images Fig. 3 Fig. 4 PMID:8692874

  8. Neuroanatomy and transgenic technologies

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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...

  9. Relative fitness of transgenic vs. non-transgenic maize x teosinte hybrids: a field evaluation.

    PubMed

    Guadagnuolo, R; Clegg, J; Ellstrand, N C

    2006-10-01

    Concern has been often expressed regarding the impact and persistence of transgenes that enter wild populations via gene flow. The impact of a transgene and its persistence are largely determined by the relative fitness of transgenic hybrids and hybrid derivatives compared to non-transgenic plants. Nevertheless, few studies have addressed this question experimentally in the field. Despite the economic importance of maize, and the fact that it naturally hybridizes with the teosinte taxon Zea mays ssp. mexicana, sometimes known as "chalco teosinte," the question has received little experimental attention in this system. Using a glyphosate-tolerant maize cultivar and chalco teosinte as parental lines, we carried out a field experiment testing (1) the relative fitness of maize x teosinte hybrids, compared to their parental taxa, as well as (2) the relative fitness of transgenic hybrids compared to non-transgenic hybrids created from the same parental stock. In order to evaluate the influence of the transgenic construct in different genetic backgrounds, our study included transgenic and non-transgenic pure maize progeny from the cultivar as well. We measured both vegetative and reproductive parameters. Our results demonstrated that hybrids have greater vigor and produced more seeds than the wild parent. However, in the absence of selective pressure from glyphosate herbicide, we did not observe any direct positive or negative impact of the transgene on the fitness or vigor of either the hybrids or pure maize progeny. We discuss our results in terms of the potential for spontaneous transgene flow and introgression from transgenic maize into sympatric teosinte.

  10. Evaluation of haematological, biochemical and histopathological parameters of transgenic rabbits.

    PubMed

    Jurcik, R; Suvegova, K; Hanusova, E; Massanyi, P; Ryban, L; Chrenek, P

    2007-11-01

    The aim of our study was to compare the hFVIII mRNA expression in different organs, pathological changes and selected haematological and biochemical blood parameters between transgenic and non-transgenic rabbits from F3 generation. Selected physiological parameters of 3- to 4-month-old transgenic rabbits from F3 generation carrying human factor VIII gene (hFVIII) were analysed and compared with those of non-transgenic ones. Before slaughtering, the blood for haematological and biochemical analysis was taken from the central ear artery. Pathological and histological examination of vital organs and RT-PCR analysis of several tissue organs of transgenic and non-transgenic animals were performed after slaughtering. Except for the mammary gland tissue, slight hfVIII mRNA expression in the spleen, lung and brain and none expression in the liver, kidney, skeletal muscle and heart of rabbits were recorded. pathological examination of vital organs showed some pathological changes in both transgenic and non-transgenic rabbits which were confirmed by histological qualitative evaluations. Statistically significant lower values of blood haemoglobin in blood of transgenic (11.86+/-0.86) animals compared with non-transgenic (12.41+/-1.02, P<0.05) ones and lower parameters of HCT (39.22+/-2.44 versus 40.89+/-2.26, P<0.01) in blood of transgenic rabbits were observed. Parameters of WBC, RBC and PLT showed no significant differences between the analysed groups. All biochemical serum parameters of transgenic rabbits were higher in comparison with non-transgenic ones. Significant differences were found in the concentration of the urea, AST and GMT between transgenic and non-transgenic animals (P<0.001) and in the total protein content, the difference was significant at P<0.05. In conclusion, our results showed that no considerable impact on the general health was found in transgenic rabbits.

  11. Improved production of genetically modified fetuses with homogeneous transgene expression after transgene integration site analysis and recloning in cattle.

    PubMed

    Bressan, Fabiana Fernandes; Dos Santos Miranda, Moyses; Perecin, Felipe; De Bem, Tiago Henrique; Pereira, Flavia Thomaz Verechia; Russo-Carbolante, Elisa Maria; Alves, Daiani; Strauss, Bryan; Bajgelman, Marcio; Krieger, José Eduardo; Binelli, Mario; Meirelles, Flavio Vieira

    2011-02-01

    Animal cloning by nuclear transfer (NT) has made the production of transgenic animals using genetically modified donor cells possible and ensures the presence of the gene construct in the offspring. The identification of transgene insertion sites in donor cells before cloning may avoid the production of animals that carry undesirable characteristics due to positional effects. This article compares blastocyst development and competence to establish pregnancies of bovine cloned embryos reconstructed with lentivirus-mediated transgenic fibroblasts containing either random integration of a transgene (random integration group) or nuclear transfer derived transgenic fibroblasts with known transgene insertion sites submitted to recloning (recloned group). In the random integration group, eGFP-expressing bovine fetal fibroblasts were selected by fluorescence activated cell sorting (FACS) and used as nuclei donor cells for NT. In the recloned group, a fibroblast cell line derived from a transgenic cloned fetus was characterized regarding transgene insertion and submitted to recloning. The recloned group had higher blastocyst production (25.38 vs. 14.42%) and higher percentage of 30-day pregnancies (14.29 vs. 2.56%) when compared to the random integration group. Relative eGFP expression analysis in fibroblasts derived from each cloned embryo revealed more homogeneous expression in the recloned group. In conclusion, the use of cell lines recovered from transgenic fetuses after identification of the transgene integration site allowed for the production of cells and fetuses with stable transgene expression, and recloning may improve transgenic animal yields.

  12. Carrying Backpacks: Physical Effects

    ERIC Educational Resources Information Center

    Illinois State Board of Education, 2006

    2006-01-01

    It is estimated that more than 40 million U.S. youth carry school materials in backs, routinely carrying books, laptop computers, personal and other items used on a daily basis. The Consumer Product Safety Commission (CPSC) estimates that 7,277 emergency visits each year result from injuries related to backpacks. Injury can occur when a child…

  13. [Transposition of the maize transposable element dSpm in transgenic sugar beets].

    PubMed

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

    2010-01-01

    Transgenic sugar beet plants carrying maize Spmn/dSpm transposable elements system have been constructed. Heterologous system of maize transposable elements Spm/dSpm was active in transgenic sugar beets that permits transposon-based gene tagging and obtaining of marker-free transgenic sugar beet.

  14. Unstable expression of transgene is associated with the methylation of CAG promoter in the offspring from the same litter of homozygous transgenic mice.

    PubMed

    Zhou, Yang; Zhang, Teng; Zhang, Qin-Kai; Jiang, Ying; Xu, Deng-Gao; Zhang, Min; Shen, Wei; Pan, Qing-Jie

    2014-08-01

    Transgenic animals have been established for studying gene function, improving animals' production traits, and providing organ models for the exploration of human diseases. However, the stability of inheritance and transgene expression in transgenic animals has gained extensive attention. The unstable expression of transgene through DNA methyltransferase (DNMT) targeting to the methylation of transgenic DNA such as CAG promoter and Egfp coding region in homozygous transgenic animals is still unknown. In the present study, the offspring from the same litter of homozygous transgenic mice carrying ubiquitously expressed enhanced green fluorescence protein driven by CMV early enhancer/chicken β-actin (CAG) promoter was observed to have unstable expression of transgene Egfp, quantitative PCR, western blot and bisulfite sequencing were conducted to quantify the expressional characteristics and methylation levels in various tissues. The correlation between transgene expression and methylation was analyzed. We have found that transgene expression is dependent on the methylation of CAG promoter, but not Egfp coding region. We have also characterized the correlation between the methylation of CAG promoter and DNMT, and found that only Dnmt3b expression is correlated with the methylation of CAG promoter. In conclusion, Dnmt3b-related methylation of CAG promoter can inhibit the transgene expression and may result in the unstable expression of transgene in the offspring from the same litter of homozygous transgenic mice.

  15. Construction of improved temperature-sensitive and mobilizable vectors and their use for constructing mutations in the adhesin-encoding acm gene of poorly transformable clinical Enterococcus faecium strains.

    PubMed

    Nallapareddy, Sreedhar R; Singh, Kavindra V; Murray, Barbara E

    2006-01-01

    Inactivation by allelic exchange in clinical isolates of the emerging nosocomial pathogen Enterococcus faecium has been hindered by lack of efficient tools, and, in this study, transformation of clinical isolates was found to be particularly problematic. For this reason, a vector for allelic replacement (pTEX5500ts) was constructed that includes (i) the pWV01-based gram-positive repAts replication region, which is known to confer a high degree of temperature intolerance, (ii) Escherichia coli oriR from pUC18, (iii) two extended multiple-cloning sites located upstream and downstream of one of the marker genes for efficient cloning of flanking regions for double-crossover mutagenesis, (iv) transcriptional terminator sites to terminate undesired readthrough, and (v) a synthetic extended promoter region containing the cat gene for allelic exchange and a high-level gentamicin resistance gene, aph(2'')-Id, to distinguish double-crossover recombination, both of which are functional in gram-positive and gram-negative backgrounds. To demonstrate the functionality of this vector, the vector was used to construct an acm (encoding an adhesin to collagen from E. faecium) deletion mutant of a poorly transformable multidrug-resistant E. faecium endocarditis isolate, TX0082. The acm-deleted strain, TX6051 (TX0082Deltaacm), was shown to lack Acm on its surface, which resulted in the abolishment of the collagen adherence phenotype observed in TX0082. A mobilizable derivative (pTEX5501ts) that contains oriT of Tn916 to facilitate conjugative transfer from the transformable E. faecalis strain JH2Sm::Tn916 to E. faecium was also constructed. Using this vector, the acm gene of a nonelectroporable E. faecium wound isolate was successfully interrupted. Thus, pTEX5500ts and its mobilizable derivative demonstrated their roles as important tools by helping to create the first reported allelic replacement in E. faecium; the constructed this acm deletion mutant will be useful for assessing the

  16. Weeding with transgenes.

    PubMed

    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.

  17. Subchronic toxicity study of GH transgenic carp.

    PubMed

    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.

  18. [Inheritance and expression stability of transgene in transgenic animals].

    PubMed

    Kong, Qing-Ran; Liu, Zhong-Hua

    2011-05-01

    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.

  19. Transgenic Crops for Herbicide Resistance

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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...

  20. Inheritance and expression of transgenes through anther culture of transgenic hot pepper.

    PubMed

    Kim, Young Soon; Kuk, Yong In; Kim, Kyung-Moon

    2007-01-01

    Anther cultures have been developed from transgenic donor peppers carrying HPT/J1-1. Eight out of sixteen plants produced from an anther culture set pepper fruits. Southern blot analysis of donors revealed two independent plants with a single copy of the integrated transgene. PCR and RT-PCR results showed the inheritance of HPT/J1-1 and expression of J1-1 in A1. All A1 progeny derived from transgenic anthers had resistance to hygromycin. They grew normally and showed similar phenotypes to the wild-type. Therefore, the use of an anther culture system coupled with genetic transformation in breeding programs will greatly facilitate the genetic improvement of pepper plants.

  1. Transgenic mammals and biotechnology.

    PubMed

    Westphal, H

    1989-02-01

    Biotechnology has begun to realize the enormous potential of transgenic technology: mice with human genes that produce human proteins of therapeutic value in their milk, pigs that express bovine genes that help them gain weight and lose backfat, animals with engineered gene defects that mimic human genetic diseases.

  2. [Progress on transgenic mosquitoes].

    PubMed

    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

  3. Transgenic Farm Animals

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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...

  4. Simple and rapid determination of homozygous transgenic mice via in vivo fluorescence imaging.

    PubMed

    Lin, Xiaolin; Jia, Junshuang; Qin, Yujuan; Lin, Xia; Li, Wei; Xiao, Gaofang; Li, Yanqing; Xie, Raoying; Huang, Hailu; Zhong, Lin; Wu, Qinghong; Wang, Wanshan; Huang, Wenhua; Yao, Kaitai; Xiao, Dong; Sun, Yan

    2015-11-17

    Setting up breeding programs for transgenic mouse strains require to distinguish homozygous from the heterozygous transgenic animals. The combinational use of the fluorescence reporter transgene and small animal in-vivo imaging system might allow us to rapidly and visually determine the transgenic mice homozygous for transgene(s) by the in vivo fluorescence imaging. RLG, RCLG or Rm17LG transgenic mice ubiquitously express red fluorescent protein (RFP). To identify homozygous RLG transgenic mice, whole-body fluorescence imaging for all of newborn F2-generation littermates produced by mating of RFP-positive heterozygous transgenic mice (F1-generation) derived from the same transgenic founder was performed. Subsequently, the immediate data analysis of the in vivo fluorescence imaging was carried out, which greatly facilitated us to rapidly and readily distinguish RLG transgenic individual(s) with strong fluorescence from the rest of F2-generation littermates, followed by further determining this/these RLG individual(s) showing strong fluorescence to be homozygous, as strongly confirmed by mouse mating. Additionally, homozygous RCLG or Rm17LG transgenic mice were also rapidly and precisely distinguished by the above-mentioned optical approach. This approach allowed us within the shortest time period to obtain 10, 8 and 2 transgenic mice homozygous for RLG, RCLG and Rm17LG transgene, respectively, as verified by mouse mating, indicating the practicality and reliability of this optical method. Taken together, our findings fully demonstrate that the in vivo fluorescence imaging offers a visual, rapid and reliable alternative method to the traditional approaches (i.e., mouse mating and real-time quantitative PCR) in identifying homozygous transgenic mice harboring fluorescence reporter transgene under the control of a ubiquitous promoter in the situation mentioned in this study.

  5. Highly Efficient Generation of Transgenic Sheep by Lentivirus Accompanying the Alteration of Methylation Status

    PubMed Central

    Liu, Chenxi; Wang, Liqin; Li, Wenrong; Zhang, Xuemei; Tian, Yongzhi; Zhang, Ning; He, Sangang; Chen, Tong; Huang, Juncheng; Liu, Mingjun

    2013-01-01

    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 genetic modification

  6. Illegal gene flow from transgenic creeping bentgrass: the saga continues.

    PubMed

    Snow, Allison A

    2012-10-01

    Ecologists have paid close attention to environmental effects that fitness-enhancing transgenes might have following crop-to-wild gene flow (e.g. Snow et al. 2003). For some crops, gene flow also can lead to legal problems,especially when government agencies have not approved transgenic events for unrestricted environmental release.Creeping bentgrass (Agrostis stolonifera), a common turf grass used in golf courses, is the focus of both areas of concern. In 2002, prior to expected deregulation (still pending), The Scotts Company planted creeping bentgrass with transgenic resistance to the herbicide glyphosate,also known as RoundUp, on 162 ha in a designated control area in central Oregon (Fig. 1).Despite efforts to restrict gene flow, wind-dispersed pollen carried transgenes to florets of local A. stolonifera and A. gigantea as far as 14 km away, and to sentinel plants placed as far as 21 km away (Watrud et al. 2004).Then, in August 2003, a strong wind event moved transgenic seeds from wind rows of cut bentgrass into nearby areas. The company’s efforts to kill all transgenic survivors in the area failed: feral glyphosate-resistant populations of A. stolonifera were found by Reichman et al.(2006), and 62% of 585 bentgrass plants had the telltale CP4 EPSPS transgene in 2006 (Zapiola et al. 2008; Fig. 2).Now, in this issue, the story gets even more interesting as Zapiola & Mallory-Smith (2012) describe a transgenic,intergeneric hybrid produced on a feral, transgenic creeping bentgrass plant that received pollen from Polypogon monspeliensis (rabbitfoot grass). Their finding raises a host of new questions about the prevalence and fitness of intergeneric hybrids, as well as how to evaluate the full extent of gene flow from transgenic crops.

  7. Relative transgene expression frequencies in homozygous versus hemizygous transgenic mice.

    PubMed

    Chang, Su-Ping; Opsahl, Margaret L; Whitelaw, C Bruce A; Morley, Steven D; West, John D

    2013-12-01

    We have used a simple binomial model of stochastic transgene inactivation at the level of the chromosome or transgene, rather than the cellular level, for the analysis of two mouse transgenic lines that show variegated patterns of expression. This predicts the percentages of cells that express one, both or neither alleles of the transgene in homozygotes from the observed percentages of cells, which express the transgene in hemizygotes. It adequately explained the relationship between the numbers of cells expressing the transgene in hemizygous and homozygous mosaic 21OH/LacZ mouse adrenals and mosaic BLG/7 mouse mammary glands. The binomial model also predicted that a small proportion of cells in mosaic mammary glands of BLG/7 homozygotes would express both BLG/7 alleles but published data indicated that all cells expressing the transgene showed monoallelic expression. Although it didn't fit all of the BLG/7 data as precisely as a more complex model, which used several ad hoc assumptions to explain these results, the simple binomial model was able to explain the relationship in observed transgene expression frequencies between hemizygous and homozygous mosaic tissues for both 21OH/LacZ and BLG/7 mice. It may prove to be a useful general model for analysing other transgenic animals showing mosaic transgene expression.

  8. Interspecies Dissemination of a Mobilizable Plasmid Harboring blaIMP-19 and the Possibility of Horizontal Gene Transfer in a Single Patient

    PubMed Central

    Gomi, Ryota; Matsuda, Tomonari; Tanaka, Michio; Nagao, Miki; Takakura, Shunji; Uemoto, Shinji; Ichiyama, Satoshi

    2016-01-01

    Carbapenemase-producing Gram-negative bacilli have been a global concern over the past 2 decades because these organisms can cause severe infections with high mortality rates. Carbapenemase genes are often carried by mobile genetic elements, and resistance plasmids can be transferred through conjugation. We conducted whole-genome sequencing (WGS) to demonstrate that the same plasmid harboring a metallo-β-lactamase gene was detected in two different species isolated from a single patient. Metallo-β-lactamase-producing Achromobacter xylosoxidans (KUN4507), non-metallo-β-lactamase-producing Klebsiella pneumoniae (KUN4843), and metallo-β-lactamase-producing K. pneumoniae (KUN5033) were sequentially isolated from a single patient and then analyzed in this study. Antimicrobial susceptibility testing, molecular typing (pulsed-field gel electrophoresis and multilocus sequence typing), and conjugation analyses were performed by conventional methods. Phylogenetic and molecular clock analysis of K. pneumoniae isolates were performed with WGS, and the nucleotide sequences of plasmids detected from these isolates were determined using WGS. Conventional molecular typing revealed that KUN4843 and KUN5033 were identical, whereas the phylogenetic tree analysis revealed a slight difference. These two isolates were separated from the most recent common ancestor 0.74 years before they were isolated. The same resistance plasmid harboring blaIMP-19 was detected in metallo-β-lactamase-producing A. xylosoxidans and K. pneumoniae. Although this plasmid was not self-transferable, the conjugation of this plasmid from A. xylosoxidans to non-metallo-β-lactamase-producing K. pneumoniae was successfully performed. The susceptibility patterns for metallo-β-lactamase-producing K. pneumoniae and the transconjugant were similar. These findings supported the possibility of the horizontal transfer of plasmid-borne blaIMP-19 from A. xylosoxidans to K. pneumoniae in a single patient. PMID

  9. Transgenes for tea?

    PubMed

    Heritage, John

    2005-01-01

    So far, no compelling scientific evidence has been found to suggest that the consumption of transgenic or genetically modified (GM) plants by animals or humans is more likely to cause harm than is the consumption of their conventional counterparts. Despite this lack of scientific evidence, the economic prospects for GM plants are probably limited in the short term and there is public opposition to the technology. Now is a good time to address several issues concerning GM plants, including the potential for transgenes to migrate from GM plants to gut microbes or to animal or human tissues, the consequences of consuming GM crops, either as fresh plants or as silage, and the problems caused by current legislation on GM labelling and beyond.

  10. Efficient and Rapid Development of Transgenic Hamster Models of TSEs Using a Radical New Technology

    DTIC Science & Technology

    2007-09-01

    high scrapie susceptibility genotype VVRRQQ. All three lines of the mouse transgenics carrying sheep, human, and elk PrP have been now re-derived. We...have observed the first transmission of the disease from our standard scrapie -infected sheep brain inoculum to the transgenic mice with sheep PrP and...already been distributed. 15. SUBJECT TERMS Transgenic, Mouse, Hamster, TSE, Sheep Scrapie , TOSK, transposon 16. SECURITY CLASSIFICATION OF

  11. Transgenics in crops

    NASA Technical Reports Server (NTRS)

    Li, Y.; Wu, Y. H.; McAvoy, R.; Duan, H.

    2001-01-01

    With rapid world population growth and declining availability of fresh water and arable land, a new technology is urgently needed to enhance agricultural productivity. Recent discoveries in the field of crop transgenics clearly demonstrate the great potential of this technology for increasing food production and improving food quality while preserving the environment for future generations. In this review, we briefly discuss some of the recent achievements in crop improvement that have been made using gene transfer technology.

  12. 35. CARRIE FURNACE No. 6 AND CAST HOUSE. THE CARRIE ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    35. CARRIE FURNACE No. 6 AND CAST HOUSE. THE CARRIE BOILER SHOP IS ON THE RIGHT, IN FRONT OF HOT BLAST STOVES. - U.S. Steel Homestead Works, Blast Furnace Plant, Along Monongahela River, Homestead, Allegheny County, PA

  13. Transgenic algae engineered for higher performance

    DOEpatents

    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.

  14. Allergenicity assessment of the Papaya ringspot virus coat protein expressed in transgenic Rainbow papaya

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The virus-resistant, transgenic commercial papaya cultivars Rainbow and SunUp (Carica papaya L.) have been consumed locally in Hawaii and elsewhere in the mainland US and Canada since their release to planters in Hawaii in 1998. These cultivars are derived from transgenic papaya line 55-1 and carry ...

  15. On fast carry select adders

    NASA Technical Reports Server (NTRS)

    Shamanna, M.; Whitaker, S.

    1992-01-01

    This paper presents an architecture for a high-speed carry select adder with very long bit lengths utilizing a conflict-free bypass scheme. The proposed scheme has almost half the number of transistors and is faster than a conventional carry select adder. A comparative study is also made between the proposed adder and a Manchester carry chain adder which shows that the proposed scheme has the same transistor count, without suffering any performance degradation, compared to the Manchester carry chain adder.

  16. On fast carry select adders

    NASA Astrophysics Data System (ADS)

    Shamanna, M.; Whitaker, S.

    This paper presents an architecture for a high-speed carry select adder with very long bit lengths utilizing a conflict-free bypass scheme. The proposed scheme has almost half the number of transistors and is faster than a conventional carry select adder. A comparative study is also made between the proposed adder and a Manchester carry chain adder which shows that the proposed scheme has the same transistor count, without suffering any performance degradation, compared to the Manchester carry chain adder.

  17. Plant biotechnology: transgenic crops.

    PubMed

    Shewry, Peter R; Jones, Huw D; Halford, Nigel G

    2008-01-01

    Transgenesis is an important adjunct to classical plant breeding, in that it allows the targeted manipulation of specific characters using genes from a range of sources. The current status of crop transformation is reviewed, including methods of gene transfer, the selection of transformed plants and control of transgene expression. The application of genetic modification technology to specific traits is then discussed, including input traits relating to crop production (herbicide tolerance and resistance to insects, pathogens and abiotic stresses) and output traits relating to the composition and quality of the harvested organs. The latter include improving the nutritional quality for consumers as well as the improvement of functional properties for food processing.

  18. An Empirical Assessment of Transgene Flow from a Bt Transgenic Poplar Plantation.

    PubMed

    Hu, Jianjun; Zhang, Jin; Chen, Xingling; Lv, Jinhui; Jia, Huixia; Zhao, Shutang; Lu, Mengzhu

    2017-01-01

    To assess the possible impact of transgenic poplar plantations on the ecosystem, we analyzed the frequency and distance of gene flow from a mature male transgenic Populus nigra plantation carrying the Bacillus thuringiensis toxin gene (Bt poplar) and the survival of Bt poplar seeds. The resultant Bt poplar seeds occurred at a frequency of ~0.15% at 0 m to ~0.02% at 500 m from the Bt poplar plantation. The germination of Bt poplar seeds diminished within three weeks in the field (germination rate from 68% to 0%) compared to 48% after three weeks of storage at 4°C. The survival rate of seedlings in the field was 0% without any treatment but increased to 1.7% under the addition of four treatments (cleaning and trimming, watering, weeding, and covering with plastic film to maintain moisture) after being seeded in the field for eight weeks. The results of this study indicate that gene flow originating from the Bt poplar plantation occurred at an extremely low level through pollen or seeds under natural conditions. This study provides first-hand field data on the extent of transgene flow in poplar plantations and offers guidance for the risk assessment of transgenic poplar plantations.

  19. An Empirical Assessment of Transgene Flow from a Bt Transgenic Poplar Plantation

    PubMed Central

    Chen, Xingling; Lv, Jinhui; Jia, Huixia; Zhao, Shutang; Lu, Mengzhu

    2017-01-01

    To assess the possible impact of transgenic poplar plantations on the ecosystem, we analyzed the frequency and distance of gene flow from a mature male transgenic Populus nigra plantation carrying the Bacillus thuringiensis toxin gene (Bt poplar) and the survival of Bt poplar seeds. The resultant Bt poplar seeds occurred at a frequency of ~0.15% at 0 m to ~0.02% at 500 m from the Bt poplar plantation. The germination of Bt poplar seeds diminished within three weeks in the field (germination rate from 68% to 0%) compared to 48% after three weeks of storage at 4°C. The survival rate of seedlings in the field was 0% without any treatment but increased to 1.7% under the addition of four treatments (cleaning and trimming, watering, weeding, and covering with plastic film to maintain moisture) after being seeded in the field for eight weeks. The results of this study indicate that gene flow originating from the Bt poplar plantation occurred at an extremely low level through pollen or seeds under natural conditions. This study provides first-hand field data on the extent of transgene flow in poplar plantations and offers guidance for the risk assessment of transgenic poplar plantations. PMID:28085955

  20. Transcriptionally Silenced Transgenes in Maize Are Activated by Three Mutations Defective in Paramutation

    PubMed Central

    McGinnis, Karen M.; Springer, Catherine; Lin, Yan; Carey, Charles C.; Chandler, Vicki

    2006-01-01

    Plants with mutations in one of three maize genes, mop1, rmr1, and rmr2, are defective in paramutation, an allele-specific interaction that leads to meiotically heritable chromatin changes. Experiments reported here demonstrate that these genes are required to maintain the transcriptional silencing of two different transgenes, suggesting that paramutation and transcriptional silencing of transgenes share mechanisms. We hypothesize that the transgenes are silenced through an RNA-directed chromatin mechanism, because mop1 encodes an RNA-dependent RNA polymerase. In all the mutants, DNA methylation was reduced in the active transgenes relative to the silent transgenes at all of the CNG sites monitored within the transgene promoter. However, asymmetrical methylation persisted at one site within the reactivated transgene in the rmr1-1 mutant. With that one mutant, rmr1-1, the transgene was efficiently resilenced upon outcrossing to reintroduce the wild-type protein. In contrast, with the mop1-1 and rmr2-1 mutants, the transgene remained active in a subset of progeny even after the wild-type proteins were reintroduced by outcrossing. Interestingly, this immunity to silencing increased as the generations progressed, consistent with a heritable chromatin state being formed at the transgene in plants carrying the mop1-1 and rmr2-1 mutations that becomes more resistant to silencing in subsequent generations. PMID:16702420

  1. Transgenic horticultural crops in Asia

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Modern biotechnology applications, including genetic engineering, are a powerful tool to complement the conventional methods of crop improvement. Asia currently has three countries cultivating biotech/transgenic crops – China, India, and the Philippines, but only China commercially grows a transgen...

  2. [Transgenics without Manichaeism].

    PubMed

    Valle, S

    2000-01-01

    We live in an era characterized by the hegemony of science and technology, an era fraught with questions awaiting answers which would enable a safe and sustainable future for humankind. The development of agro-industrial processes - food products in particular - through recombinant DNA technology has enhanced the profit prospects of the few big biotechnology companies and of large-scale farmers who have access to the latest technological developments. We thus oppose a moratorium on recombinant DNA technology. Moreover, hasty statements about risk-free transgenics may be misleading in the absence of extensive safety tests. There is a pressing need for the establishment of biosafety policy in this country involving the organized civil society and every government agency responsible for monitoring such matters. There is also the need to put in place a bio-surveillance and a code of ethics regarding genetic manipulation.

  3. Superoxide dismutase transgenes in sugarbeets confer resistance to oxidative agents and the fungus C. beticola.

    PubMed

    Tertivanidis, Konstantinos; Goudoula, Catherine; Vasilikiotis, Christos; Hassiotou, Efthymia; Perl-Treves, Rafael; Tsaftaris, Athanasios

    2004-06-01

    Sugarbeets carrying superoxide dismutase transgenes were developed in order to investigate the possibility of enhancing their resistance to oxidative stress. Binary T-DNA vectors carrying the chloroplastic and cytosolic superoxide dismutase genes from tomato, were used for Agrobacterium-mediated transformation of sugarbeet petioles. The transgenic plants were subjected to treatments known to cause oxidative stress, such as the herbicide methyl viologen and a natural photosensitizer toxin produced by the fungus Cercospora beticola, namely cercosporin. The transgenic plants exhibited increased tolerance to methyl viologen, to pure cercosporin, as well as to leaf infection with the fungus C. beticola.

  4. Creation of transgenic rice plants producing small interfering RNA of Rice tungro spherical virus.

    PubMed

    Le, Dung Tien; Chu, Ha Duc; Sasaya, Takahide

    2015-01-01

    Rice tungro spherical virus (RTSV), also known as Rice waika virus, does not cause visible symptoms in infected rice plants. However, the virus plays a critical role in spreading Rice tungro bacilliform virus (RTBV), which is the major cause of severe symptoms of rice tungro disease. Recent studies showed that RNA interference (RNAi) can be used to develop virus-resistance transgenic rice plants. In this report, we presented simple procedures and protocols needed for the creation of transgenic rice plants capable of producing small interfering RNA specific against RTSV sequences. Notably, our study showed that 60 out of 64 individual hygromycin-resistant lines (putative transgenic lines) obtained through transformation carried transgenes designed for producing hairpin double-stranded RNA. Northern blot analyses revealed the presence of small interfering RNA of 21- to 24-mer in 46 out of 56 confirmed transgenic lines. Taken together, our study indicated that transgenic rice plants carrying an inverted repeat of 500-bp fragments encoding various proteins of RTSV can produce small interfering RNA from the hairpin RNA transcribed from that transgene. In light of recent studies with other viruses, it is possible that some of these transgenic rice lines might be resistant to RTSV.

  5. Transgenic mice in developmental toxicology

    SciTech Connect

    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.

  6. Transgenic mice in developmental toxicology

    SciTech Connect

    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.

  7. Transgenic, inducible RNAi in megakaryocytes and platelets in mice

    PubMed Central

    TAKIGUCHI, M.; JAMES, C.; JOSEFSSON, E. C.; CARMICHAEL, C. L.; PREMSRIRUT, P. K.; LOWE, S. W.; HAMILTON, J. R.; HUANG, D. C. S.; KILE, B. T.; DICKINS, R. A.

    2012-01-01

    Summary Background RNA interference (RNAi) is a powerful tool for suppressing gene function. The tetracycline (tet)-regulated expression system has recently been adapted to allow inducible RNAi in mice, however its efficiency in a particular cell type in vivo depends on a transgenic tet transactivator expression pattern and is often highly variable. Objective We aimed to establish a transgenic strategy that allows efficient and inducible gene knockdown in particular hematopoietic lineages in mice. Methods and results Using a tet-regulated reporter gene strategy, we found that transgenic mice expressing the rtTA (tet-on) transactivator under control of the cytomegalovirus (CMV) promoter (CMV-rtTA mice) display inducible reporter gene expression with unusual and near-complete efficiency in megakaryocytes and platelets. To test whether the CMV-rtTA transgene can drive inducible and efficient gene knockdown within this lineage, we generated a novel mouse strain harboring a tet-regulated short hairpin RNA (shRNA) targeting Bcl-xL, a pro-survival Bcl-2 family member known to be essential for maintaining platelet survival. Doxycycline treatment of adult mice carrying both transgenes induces shRNA expression, depletes Bcl-xL in megakaryocytes and triggers severe thrombocytopenia, whereas doxycycline withdrawal shuts off shRNA expression, normalizes Bcl-xL levels and restores platelet numbers. These effects are akin to those observed with drugs that target Bcl-xL, clearly demonstrating that this transgenic system allows efficient and inducible inhibition of genes in megakaryocytes and platelets. Conclusions We have established a novel transgenic strategy for inducible gene knockdown inmegakaryocytes and platelets that will be useful for characterizing genes involved in platelet production and function in adult mice. PMID:21138522

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

    PubMed

    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

  9. Generation of domestic transgenic cloned kittens using lentivirus vectors.

    PubMed

    Gómez, Martha C; Pope, Charles Earle; Kutner, Robert H; Ricks, David M; Lyons, Leslie A; Ruhe, Mark T; Dumas, Cherie; Lyons, Justine; Dresser, Betsy L; Reiser, Jakob

    2009-03-01

    The efficient use of somatic cell nuclear transfer (SCNT), in conjunction with genetic modification of donor cells provides a general means to add or inactivate genes in mammals. This strategy has substantially improved the efficacy of producing genetically identical animals carrying mutant genes corresponding to specific human disorders. Lentiviral (LV) vectors have been shown to be well suited for introducing transgenes into cells to be used as donor nuclei for SCNT. In the present study, we established an LV vector-based transgene delivery approach for producing live transgenic domestic cats by SCNT. We have demonstrated that cat fetal fibroblasts can be transduced with EGFP-encoding LV vectors bearing various promoters including the human cytomegalovirus immediate early (hCMV-IE) promoter, the human translation elongation factor 1alpha (hEF-1alpha) promoter and the human ubiquitin C (hUbC) promoter. Among the promoters tested, embryos reconstructed with donor cells transduced with a LV-vector bearing the hUbC promoter displayed sustained transgene expression at the blastocyst stage while embryos reconstructed with LV vector-transduced cells containing hCMV-IE-EGFP or hEF-1alpha-EGFP cassettes did not. After transfer of 291 transgenic cloned embryos into the oviducts of eight recipient domestic cats (mean =36.5 +/- 10.1), three (37.5%) were diagnosed to be pregnant, and a total of six embryos (2.1%) implanted. One live male offspring was delivered by Cesarean section on day 64 of gestation, and two kittens were born dead after premature delivery on day 55. In summary, we report the birth of transgenic cloned kittens produced by LV vector-mediated transduction of donor cells and confirm that cloned kittens express the EGFP reporter transgene in all body tissues.

  10. Transgenic plants for phytoremediation.

    PubMed

    Maestri, Elena; Marmiroli, Nelson

    2011-01-01

    Phytoremediation is a green, sustainable and promising solution to problems of environmental contamination. It entails the use of plants for uptake, sequestration, detoxification or volatilization of inorganic and organic pollutants from soils, water, sediments and possibly air. Phytoremediation was born from the observation that plants possessed physiological properties useful for environmental remediation. This was shortly followed by the application of breeding techniques and artificial selection to genetically improve some of the more promising and interesting species. Now, after nearly 20 years of research, transgenic plants for phytoremediation have been produced, but none have reached commercial existence. Three main approaches have been developed: (1) transformation with genes from other organisms (mammals, bacteria, etc.); (2) transformation with genes from other plant species; and (3) overexpression of genes from the same plant species. Many encouraging results have been reported, even though in some instances results have been contrary to expectations. This review will illustrate the main examples with a critical discussion of what we have learnt from them.

  11. Assessment of the diversity and dynamics of Plum pox virus and aphid populations in transgenic European plums under Mediterranean conditions.

    PubMed

    Capote, Nieves; Pérez-Panadés, Jordi; Monzó, César; Carbonell, Emilio; Urbaneja, Alberto; Scorza, Ralph; Ravelonandro, Michel; Cambra, Mariano

    2008-06-01

    The molecular variability of Plum pox virus (PPV) populations was compared in transgenic European plums (Prunus domestica L.) carrying the coat protein (CP) gene of PPV and non-transgenic plums in an experimental orchard in Valencia, Spain. A major objective of this study was to detect recombination between PPV CP transgene transcripts and infecting PPV RNA. Additionally, we assessed the number and species of PPV aphid vectors that visited transgenic and non-transgenic plum trees. Test trees consisted of five different P. domestica transgenic lines, i.e. the PPV-resistant C5 'HoneySweet' line and the PPV-susceptible C4, C6, PT6 and PT23 lines, and non-transgenic P. domestica and P. salicina Lind trees. No significant difference in the genetic diversity of PPV populations infecting transgenic and conventional plums was detected, in particular no recombinant between transgene transcripts and incoming viral RNA was found at detectable levels. Also, no significant difference was detected in aphid populations, including viruliferous individuals, that visited transgenic and conventional plums. Our data indicate that PPV-CP transgenic European plums exposed to natural PPV infection over an 8 year period caused limited, if any, risk beyond the cultivation of conventional plums under Mediterranean conditions in terms of the emergence of recombinant PPV and diversity of PPV and aphid populations.

  12. Transgenic Gladiolus plants transformed with the bean yellow mosaic virus coat-protein gene in either sense or antisense orientation.

    PubMed

    Kamo, Kathryn; Gera, Abed; Cohen, Jacob; Hammond, John; Blowers, Alan; Smith, Franzine; Van Eck, Joyce

    2005-02-01

    Transgenic Gladiolus plants transformed with the bean yellow mosaic virus (BYMV) coat-protein (CP) gene in either sense or antisense (AS) orientation were developed using biolistics. Four of the plants were confirmed to carry the CP gene in the sense orientation of the gene and seven plants in the AS orientation. Two of the CP plant lines and all of the AS lines showed DNA rearrangements of the transgene in addition to an intact copy of the transgene. The copy number ranged from one to nine. Of the 11 lines, eight had only one to four copies of the transgene. Transcription of the transgene occurred for three of the CP lines and five of the AS lines as determined by Northern hybridization. All 11 plant lines were challenged with BYMV using controlled aphid transmission. One month following aphid transmission, the transgenic plants were examined by immunoelectron microscopy for presence of the virus. Several transgenic plant lines containing either antiviral transgene showed a lower incidence of infection (percentage of plants infected as detected by immunoelectron microscopy) than the non-transformed plants. Most of the CP- and AS-transgenic plants that did not contain BYMV 1 month after challenge were found to contain BYMV the next season. It appeared that BYMV infection was delayed in the CP- and AS-transgenic lines but that the transgenes did not prevent eventual infection of BYMV. This is the first report of developing a floral bulb crop with antiviral genes to BYMV.

  13. Oat Phytochrome Is Biologically Active in Transgenic Tomatoes.

    PubMed

    Boylan, M. T.; Quail, P. H.

    1989-08-01

    To determine the functional homology between phytochromes from evolutionarily divergent species, we used the cauliflower mosaic virus 35S promoter to express a monocot (oat) phytochrome cDNA in a dicot plant (tomato). Immunoblot analysis shows that more than 50% of the transgenic tomato plants synthesize the full-length oat phytochrome polypeptide. Moreover, leaves of light-grown transgenic plants contain appreciably less oat phytochrome than leaves from dark-adapted plants, and etiolated R1 transgenic seedlings have higher levels of spectrally active phytochrome than wild-type tomato seedlings in direct proportion to the level of immunochemically detectable oat polypeptide present. These data suggest that the heterologous oat polypeptide carries a functional chromophore, allowing reversible photoconversion between the two forms of the molecule, and that the far-red absorbing form (Pfr) is recognized and selectively degraded by the Pfr-specific degradative machinery in the dicot cell. The overexpression of oat phytochrome has pleiotropic, phenotypic consequences at all major phases of the life cycle. Adult transgenic tomato plants expressing high levels of the oat protein tend to be dwarfed, with dark green foliage and fruits. R1 transgenic seedlings have short hypocotyls with elevated anthocyanin contents. We conclude that a monocot phytochrome can be synthesized and correctly processed to a biologically active form in a dicot cell, and that the transduction pathway components that interact with the photoreceptor are evolutionarily conserved.

  14. Fitness of anopheline mosquitoes expressing transgenes that inhibit Plasmodium development.

    PubMed

    Moreira, Luciano A; Wang, Jing; Collins, Frank H; Jacobs-Lorena, Marcelo

    2004-03-01

    One potential strategy for the control of malaria and other vector-borne diseases is the introduction into wild vector populations of genetic constructs that reduce vectorial capacity. An important caveat of this approach is that the genetic construct should have minimal fitness cost to the transformed vector. Previously, we produced transgenic Anopheles stephensi expressing either of two effector genes, a tetramer of the SM1 dodecapeptide or the phospholipase A2 gene (PLA2) from honeybee venom. Mosquitoes carrying either of these transgenes were impaired for Plasmodium berghei transmission. We have investigated the role of two effector genes for malaria parasite blockage in terms of the fitness imposed to the mosquito vector that expresses either molecule. By measuring mosquito survival, fecundity, fertility, and by running population cage experiments, we found that mosquitoes transformed with the SM1 construct showed no significant reduction in these fitness parameters relative to nontransgenic controls. The PLA2 transgenics, however, had reduced fitness that seemed to be independent of the insertion site of the transgene. We conclude that the fitness load imposed by refractory gene(s)-expressing mosquitoes depends on the effect of the transgenic protein produced in that mosquito. These results have important implications for implementation of malaria control via genetic modification of mosquitoes.

  15. Carry Groups: Abstract Algebra Projects

    ERIC Educational Resources Information Center

    Miller, Cheryl Chute; Madore, Blair F.

    2004-01-01

    Carry Groups are a wonderful collection of groups to introduce in an undergraduate Abstract Algebra course. These groups are straightforward to define but have interesting structures for students to discover. We describe these groups and give examples of in-class group projects that were developed and used by Miller.

  16. Suppression of intestinal immunity through silencing of TCTP by RNAi in transgenic silkworm, Bombyx mori.

    PubMed

    Hu, Cuimei; Wang, Fei; Ma, Sanyuan; Li, Xianyang; Song, Liang; Hua, Xiaoting; Xia, Qingyou

    2015-12-10

    Intestinal immune response is a front line of host defense. The host factors that participate in intestinal immunity response remain largely unknown. We recently reported that Translationally Controlled Tumor Protein (BmTCTP) was obtained by constructing a phage display cDNA library of the silkworm midgut and carrying out high throughput screening of pathogen binding molecules. To further address the function of BmTCTP in silkworm intestinal immunity, transgenic RNAi silkworms were constructed by microinjection piggBac plasmid to Dazao embryos. The antimicrobial capacity of transgenic silkworm decreased since the expression of gut antimicrobial peptide from transgenic silkworm was not sufficiently induced during oral microbial challenge. Moreover, dynamic ERK phosphorylation from transgenic silkworm midgut was disrupted. Taken together, the innate immunity of intestinal was suppressed through disruption of dynamic ERK phosphorylation after oral microbial infection as a result of RNAi-mediated knockdown of midgut TCTP in transgenic silkworm.

  17. Analysis of transgenic wheat (Triticum aestivum L.) harboring a maize (Zea mays L.) gene for plastid EF-Tu: segregation pattern, expression and effects of the transgene.

    PubMed

    Fu, Jianming; Ristic, Zoran

    2010-06-01

    We previously reported that transgenic wheat (Triticum aestivum L.) carrying a maize (Zea mays L.) gene (Zmeftu1) for chloroplast protein synthesis elongation factor, EF-Tu, displays reduced thermal aggregation of leaf proteins, reduced injury to photosynthetic membranes (thylakoids), and enhanced rate of CO(2) fixation following exposure to heat stress (18 h at 45 degrees C) [Fu et al. in Plant Mol Biol 68:277-288, 2008]. In the current study, we investigated the segregation pattern and expression of the transgene Zmeftu1 and determined the grain yield of transgenic plants after exposure to a brief heat stress (18 h at 45 degrees C). We also assessed thermal aggregation of soluble leaf proteins in transgenic plants, testing the hypothesis that increased levels of EF-Tu will lead to a non-specific protection of leaf proteins against thermal aggregation. The transgenic wheat displayed a single-gene pattern of segregation of Zmeftu1. Zmeftu1 was expressed, and the transgenic plants synthesized and accumulated three anti-EF-Tu cross-reacting polypeptides of similar molecular mass but different pI, suggesting the possibility of posttranslational modification of this protein. The transgenic plants also showed better grain yield after exposure to heat stress compared with their non-transgenic counterparts. Soluble leaf proteins of various molecular masses displayed lower thermal aggregation in transgenic than in non-transgenic wheat. The results suggest that overexpression of chloroplast EF-Tu can be beneficial to wheat tolerance to heat stress. Moreover, the results also support the hypothesis that EF-Tu contributes to heat tolerance by acting as a molecular chaperone and protecting heat-labile proteins from thermal aggregation in a non-specific manner.

  18. The IncP-6 Plasmid p10265-KPC from Pseudomonas aeruginosa Carries a Novel ΔISEc33-Associated blaKPC-2 Gene Cluster

    PubMed Central

    Dai, Xiaotian; Zhou, Dongsheng; Xiong, Wei; Feng, Jiao; Luo, Wenbo; Luo, Guangming; Wang, Haijing; Sun, Fengjun; Zhou, Xiangdong

    2016-01-01

    Pseudomonas aeruginosa strain 10265 was recovered from a patient with pneumonia in a Chinese public hospital, and it displays the carbapenem resistance phenotype due to the acquisition of a non-conjugative but mobilizable IncP-6-type plasmid p10265-KPC. p10265-KPC carries a Tn5563-borne defective mer locus, and a novel ΔISEc33-associated blaKPC-2 gene cluster without paired inverted repeats and paired direct repeats at both ends. Mobilization of this ΔISEc33-associated element in p10265-KPC would be attributed to homologous recombination-based insertion of a foreign structure Tn3-ISApu1-orf7-ISApu2- ISKpn27-ΔblaTEM-1-blaKPC-2-ΔISKpn6- korC-orf6-klcA-ΔrepB into a pre-existent intact ISEc33, making ISEc33 truncated at the 3′ end. The previously reported pCOL-1 represents the first sequenced KPC-producing IncP-6 plasmid, while p10265-KPC is the second one. These two plasmids carry two distinct blaKPC-2 gene clusters, which are inserted into the different sites of the IncP-6 backbone and have different evolutionary histories of assembly and mobilization. This is the first report of identification of the IncP-6-type resistance plasmid in China. PMID:27014233

  19. The IncP-6 Plasmid p10265-KPC from Pseudomonas aeruginosa Carries a Novel ΔISEc33-Associated bla KPC-2 Gene Cluster.

    PubMed

    Dai, Xiaotian; Zhou, Dongsheng; Xiong, Wei; Feng, Jiao; Luo, Wenbo; Luo, Guangming; Wang, Haijing; Sun, Fengjun; Zhou, Xiangdong

    2016-01-01

    Pseudomonas aeruginosa strain 10265 was recovered from a patient with pneumonia in a Chinese public hospital, and it displays the carbapenem resistance phenotype due to the acquisition of a non-conjugative but mobilizable IncP-6-type plasmid p10265-KPC. p10265-KPC carries a Tn5563-borne defective mer locus, and a novel ΔISEc33-associated bla KPC-2 gene cluster without paired inverted repeats and paired direct repeats at both ends. Mobilization of this ΔISEc33-associated element in p10265-KPC would be attributed to homologous recombination-based insertion of a foreign structure Tn3-ISApu1-orf7-ISApu2- ISKpn27-Δbla TEM-1 -bla KPC-2 -ΔISKpn6- korC-orf6-klcA-ΔrepB into a pre-existent intact ISEc33, making ISEc33 truncated at the 3' end. The previously reported pCOL-1 represents the first sequenced KPC-producing IncP-6 plasmid, while p10265-KPC is the second one. These two plasmids carry two distinct bla KPC-2 gene clusters, which are inserted into the different sites of the IncP-6 backbone and have different evolutionary histories of assembly and mobilization. This is the first report of identification of the IncP-6-type resistance plasmid in China.

  20. Phenotypic performance of transgenic potato (Solanum tuberosum L.) plants with pyramided rice cystatin genes (OCI and OCII)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The evaluation of transgenic plants commonly carried out under controlled conditions in culture rooms and greenhouses can give valuable information about the influence of introduced genes on transgenic plant phenotype. However, an overall assessment of plant performance can only be made by testing t...

  1. Metabolite fingerprinting in transgenic lettuce.

    PubMed

    Garratt, Lee C; Linforth, Robert; Taylor, Andrew J; Lowe, Kenneth C; Power, J Brian; Davey, Michael R

    2005-03-01

    Metabolite fingerprinting has been achieved using direct atmospheric pressure chemical ionization-mass spectrometry (APCI-MS) and linked gas chromatography (GC-APCI/EI-MS) for transgenic lettuce (Lactuca sativa L. cv. Evola) plants expressing an IPT gene under the control of the senescence-specific SAG12 promoter from Arabidopsis thaliana (P(SAG12)-IPT). Mature heads of transgenic lettuce and their azygous controls were maintained under defined conditions to assess their shelf life. Transgenic lettuce plants exhibited delayed senescence and significant increases (up to a maximum of threefold) in the concentrations of three volatile organic compounds (VOCs), corresponding to molecular masses of 45, 47 and 63, when compared with heads from azygous plants. These VOCs were identified as acetaldehyde (45), ethanol (47) and dimethyl sulphide (63). The increase in dimethyl sulphide was paralleled by an accumulation of reactive oxygen species (ROS) in the heads of transgenic plants. These results demonstrate the applicability of metabolic fingerprinting techniques to elucidate the underlying pleiotropic responses of plants to transgene expression.

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

    PubMed

    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.

  3. Expression specificity of GFAP transgenes.

    PubMed

    Su, Mu; Hu, Huimin; Lee, Youngjin; d'Azzo, Alessandra; Messing, Albee; Brenner, Michael

    2004-11-01

    Glial fibrillary acidic protein (GFAP) is an intermediate filament protein found predominantly in astrocytes. This specificity has recommended the GFAP gene promoter for targeting transgene expression to astrocytes. Although both we [Brenner et al. J. Neurosci. 14:1030-1037, (1994)] and others [Mucke et al. New Biol. 3:465-474, (1991)] have reported astrocyte specificity for GFAP promoters, we demonstrate here that these DNA sequences can also direct activity in neurons. The pattern of neuronal activity varied with both the nature of the expressed sequence and the transgene insertion site. Specifically, neuronal expression was very high for a protective protein/cathepsin A minigene, moderate for lacZ and undetectable for GFP. These findings, coupled with a survey of the literature, recommend that investigators using GFAP-driven transgenes verify specificity for each line studied, using a detection system whose sensitivity is sufficient to detect a compromising level of misexpression.

  4. Comparative persistence of thiacloprid in Bt-transgenic cabbage (Brassica oleracea cv. capitata) vis-à-vis non-transgenic crop and its decontamination.

    PubMed

    Dutta, Debashis; Niwas, Ram; Gopal, Madhuban

    2012-11-01

    Thiacloprid is a systemic neonicotinoid. The study hypothesized that difference may be seen in the rate of dissipation of thiacloprid when applied on non-transgenic and transgenic cabbage. Thiacloprid was estimated by HPLC. Half life of thiacloprid in transgenic as well as in normal cabbage ranged between 12.3-13.1 days in two doses of application. Under field condition, after 15 days, 59.2% and 54.3% dissipation was recorded at lower and higher rates of application in transgenic cabbage, where as the insecticide dissipated 57.5% and 59.1% for single dose and double dose application, respectively in non-transgenic cabbage. The study establishes that there is no significant difference in dissipation of a systemic pesticide in transgenic versus non-transgenic cabbage. Decontamination of thiacloprid contaminated cabbage was carried out by different chemical treatments. The application of 0.5% NaHCO(3) (an edible alkali) may be recommended for decontamination. Thiacloprid residues in the day-3 field samples of cabbage could be reduced below Japanese MRL (1.0 mg kg(-1)) by treating with 0.5% NaHCO(3) solution for 1 h.

  5. Molecular control of transgene escape from genetically modified plants.

    PubMed

    Kuvshinov, V; Koivu, K; Kanerva, A; Pehu, E

    2001-02-05

    Potential risks of gene escape from transgenic crops through pollen and seed dispersal are being actively discussed and have slowed down full utilization of gene technology in crop improvement. To ban the transgene flow, barren zones and 'terminator' technology were developed as GMO risk management technologies in transgenic crops. Unfortunately, the technologies have not protected reliably the transgene migration to wild relatives. The present study offers a novel molecular technique to eliminate gene flow from transgenic plants to wild relatives by recoverable block of function (RBF). The RBF consists of a blocking sequence linked to the gene of interest and a recovering sequence, all in one transformable construct. The blocking sequence blocks a certain molecular or physiological function of the host plant. Action of the blocking sequence leads to the death of the host plant or to an alteration in its phenotype resulting in inability for sexual reproduction in nature. The recovering construct recovers the blocked function of the host plant. The recovering construct is regulated externally by a specific chemical or physical treatment of the plants and does not act under natural conditions. In nature, hybrids of the transgenic plants with its wild relatives carrying the RBF will die or be unable to reproduce because of the blocking construct action. A working model of RBF is described in this report as one example of the RBF concept. This RBF example is based on barnase (the blocking construct) and barstar (the recovering construct) gene expression in tobacco under sulfhydryl endopeptidase (SH-EP) and a heat shock (HS) promoter, respectively.

  6. [The exploration and practice of production of transgenic zebrafish into undergraduate student gene engineering experimental teaching].

    PubMed

    Yuan, Wu-Zhou; Deng, Yun

    2013-11-01

    The preparation of transgenic animals is one of the core technology and critical achievement of gene engineering. However, it has not been reported that the gene engineering experimental course of undergraduate students in universities of mainland China has carried out the preparation of transgenic animals. In this paper, the authors took the advantage of scientific research platform, introduced the transgenic zebrafish technology to gene engineering experimental course of undergraduate students, and explored and practiced related teaching model, which had achieved good results and had great value to popularize.

  7. Fructan biosynthesis in transgenic plants.

    PubMed

    Cairns, Andrew J

    2003-01-01

    Data from plants transformed to accumulate fructan are assessed in the context of natural concentrations of reserve carbohydrates and natural fluxes of carbon in primary metabolism: Transgenic fructan accumulation is universally reported as an instantaneous endpoint concentration. In exceptional cases, concentrations of 60-160 mg g(-1) fresh mass were reported and compare favourably with naturally occurring maximal starch and fructan content in leaves and storage organs. Generally, values were less than 20 mg g(-1) for plants transformed with bacterial genes and <9 mg g(-1) for plant-plant transformants. Superficially, the results indicate a marked modification of carbon partitioning. However, transgenic fructan accumulation was generally constitutive and involved accumulation over time-scales of weeks or months. When calculated as a function of accumulation period, fluxes into the transgenic product were low, in the range 0.00002-0.03 nkat g(-1). By comparison with an estimated minimum daily carbohydrate flux in leaves for a natural fructan-accumulating plant in field conditions (37 nkat g(-1)), transgenic fructan accumulation was only 0.00005-0.08% of primary carbohydrate flux and does not indicate radical modification of carbon partitioning, but rather, a quantitatively minor leakage into transgenic fructan. Possible mechanisms for this low fructan accumulation in the transformants are considered and include: (i) rare codon usage in bacterial genes compared with eukaryotes, (ii) low transgene mRNA concentrations caused by low expression and/or high turnover, (iii) resultant low expression of enzyme protein, (iv) resultant low total enzyme activity, (v) inappropriate kinetic properties of the gene products with respect to substrate concentrations in the host, (vi) in situ product hydrolysis, and (vii) levan toxicity. Transformants expressing bacterial fructan synthesis exhibited a number of aberrant phenotypes such as stunting, leaf bleaching, necrosis, reduced

  8. How To Produce and Characterize Transgenic Plants.

    ERIC Educational Resources Information Center

    Savka, Michael A.; Wang, Shu-Yi; Wilson, Mark

    2002-01-01

    Explains the process of establishing transgenic plants which is a very important tool in plant biology and modern agriculture. Produces transgenic plants with the ability to synthesize opines. (Contains 17 references.) (YDS)

  9. Immunological Prevention of Spontaneous Mammary Carcinoma in Transgenic Mice

    DTIC Science & Technology

    2001-08-01

    developed more slowly by transgenic FVB Anatomia Patologica, Ospedale S.S. Annunziata, Via Valignani, 66100 female mice carrying the wild-type proto...coopted (Pezzella et al., 1997). Anatomia Patologica. Ospedale SS. Annunziata, Via Valignani, 66100 Chieti, Italy. Fax: 39 0871 330471. E-mail: musiani...lo Studio e la Cura dei Tumori, Milan, Italy; and Reprints: Piero Musiani, G. d’ Annunzio University of Chieti, Anatomia Department of Experimental

  10. Why do dolphins carry sponges?

    PubMed

    Mann, Janet; Sargeant, Brooke L; Watson-Capps, Jana J; Gibson, Quincy A; Heithaus, Michael R; Connor, Richard C; Patterson, Eric

    2008-01-01

    Tool use is rare in wild animals, but of widespread interest because of its relationship to animal cognition, social learning and culture. Despite such attention, quantifying the costs and benefits of tool use has been difficult, largely because if tool use occurs, all population members typically exhibit the behavior. In Shark Bay, Australia, only a subset of the bottlenose dolphin population uses marine sponges as tools, providing an opportunity to assess both proximate and ultimate costs and benefits and document patterns of transmission. We compared sponge-carrying (sponger) females to non-sponge-carrying (non-sponger) females and show that spongers were more solitary, spent more time in deep water channel habitats, dived for longer durations, and devoted more time to foraging than non-spongers; and, even with these potential proximate costs, calving success of sponger females was not significantly different from non-spongers. We also show a clear female-bias in the ontogeny of sponging. With a solitary lifestyle, specialization, and high foraging demands, spongers used tools more than any non-human animal. We suggest that the ecological, social, and developmental mechanisms involved likely (1) help explain the high intrapopulation variation in female behaviour, (2) indicate tradeoffs (e.g., time allocation) between ecological and social factors and, (3) constrain the spread of this innovation to primarily vertical transmission.

  11. Human health and transgenic crops

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Under the joint auspices of the Agrochemical and the Agricultural and Food Chemistry Divisions of the American Chemical Society, we organized a short symposium on “Human Health and Transgenic Crops” at the 244th ACS national meeting, held August 19-23, 2012 in Philadelphia, PA, to examine an array o...

  12. Novelties that change carrying capacity.

    PubMed

    Erwin, Douglas H

    2012-09-01

    Comparative developmental studies have revealed a rich array of details about the patterns and processes of morphological change in animals and increasingly in plants. But, applying these insights to the study of major episodes of evolutionary innovation requires understanding how these novel morphologies become established and sufficiently abundant (either as individuals within a species or as a clade of species) to be preserved in the fossil record, and, in many cases, to influence ecological processes. Evolutionary novelties may: (1) disappear without changing the species; (2) be associated with the generation (through selection or drift) of a new species; and if the latter (3) may or may not become ecologically significant. Only the latter are commonly preserved in the fossil record. These alternatives mirror the distinction among historians of technology between innovation and invention. Here, I argue that specific sorts of evolutionary inventions drive ecological transformation, essentially constructing an environment for themselves and ancillary organisms through ecological spillover effects, increasing the "carrying capacity" of an ecosystem.

  13. Carrying our founders' mission overseas.

    PubMed

    Williams, Patricia A

    2006-01-01

    Catholic' health care providers have a calling to care for people in need, and that mission does not stop at geographical boundaries. In fact, U.S. health facilities in many cases were founded by overseas religious communities with a mission. Providing aid internationally enables U.S. sites to carry on that legacy. Although Americans traveling overseas to provide aid usually expect to be "teachers", they often find themselves becoming "students" instead. They learn to provide care without the advanced technology that is available in developed countries. They often experience cultures in which people can only hope for care access and in which patients are deeply appreciative of the services they receive. This type of education can change U.S. health care providers' perspective of their role and of the services they deliver. While gaining this wisdom-and imparting their own knowledge-providers also affect the quality of life of people in developing countries. In the end, global aid can create a better world for everyone, benefiting not only the recipients but also the worldwide community. When developing countries become more stable, develop stronger infrastructures, and have healthier citizens, other countries benefit from this progress.

  14. Detecting and quantifying the adventitious presence of transgenic seeds in safflower, Carthamus tinctorius L.

    PubMed

    Christianson, Jed; McPherson, Marc; Topinka, Deborah; Hall, Linda; Good, Allen G

    2008-07-23

    Safflower ( Carthamus tinctorius L.) is currently being developed as a platform for the production of novel proteins. Methods for detecting and quantifying transgenic safflower are needed to ensure seed quality and to monitor for its adventitious presence. We developed and compared three methods of assaying for transgenic safflower presence in conventional seedlots: field bioassays, enzyme-linked immunosorbent assays (ELISA), and quantitative polymerase chain reaction (Q-PCR). Limits for reliable quantification for both ELISA and Q-PCR are approximately 0.1%, although levels at least as low as 0.02% can be detected by Q-PCR. Levels of quantification for the field bioassay are limited only by space and resources available. Multiple sampling methods to detect and quantify transgenic safflower presence at levels lower than 0.1% were used on field collected samples from a pollen outcrossing experiment to quantify the adventitious presence of transgenic safflower. Taking into account the potential utility and relative advantages or disadvantages of each detection method, it is recommended that the initial testing for the adventitious presence of transgenic seed be carried out using an antibody-based test if available and that Q-PCR-based assays to quantify transgenic proportion be used when it is necessary to identify specific transgenic constructs or if antibody-based assays are not readily available.

  15. Breeding of transgenic cattle for human coagulation factor IX by a combination of lentiviral system and cloning.

    PubMed

    Monzani, P S; Sangalli, J R; De Bem, T H C; Bressan, F F; Fantinato-Neto, P; Pimentel, J R V; Birgel-Junior, E H; Fontes, A M; Covas, D T; Meirelles, F V

    2013-02-28

    Recombinant coagulation factor IX must be produced in mammalian cells because FIX synthesis involves translational modifications. Human cell culture-based expression of human coagulation factor IX (hFIX) is expensive, and large-scale production capacity is limited. Transgenic animals may greatly increase the yield of therapeutic proteins and reduce costs. In this study, we used a lentiviral system to obtain transgenic cells and somatic cell nuclear transfer (SCNT) to produce transgenic animals. Lentiviral vectors carrying hFIX driven by 3 bovine β-casein promoters were constructed. Bovine epithelial mammary cells were transduced by lentivirus, selected with blasticidin, plated on extracellular matrix, and induced by lactogenic hormones; promoter activity was evaluated by quantitative PCR. Transcriptional activity of the 5.335-kb promoter was 6-fold higher than the 3.392- and 4.279-kb promoters, which did not significantly differ. Transgenic bovine fibroblasts were transduced with lentivirus carrying the 5.335-kb promoter and used as donor cells for SCNT. Cloned transgenic embryo production yielded development rates of 28.4%, similar to previous reports on cloned non-transgenic embryos. The embryos were transferred to recipient cows (N = 21) and 2 births of cloned transgenic cattle were obtained. These results suggest combination of the lentiviral system and cloning may be a good strategy for production of transgenic cattle.

  16. Sequence homology requirements for transcriptional silencing of 35S transgenes and post-transcriptional silencing of nitrite reductase (trans)genes by the tobacco 271 locus.

    PubMed

    Thierry, D; Vaucheret, H

    1996-12-01

    The transgene locus of the tobacco plant 271 (271 locus) is located on a telomere and consists of multiple copies of a plasmid carrying an NptII marker gene driven by the cauliflower mosaic virus (CaMV) 19S promoter and the leaf-specific nitrite reductase Nii1 cDNA cloned in the antisense orientation under the control of the CaMV 35S promoter. Previous analysis of gene expression in leaves has shown that this locus triggers both post-transcriptional silencing of the host leaf-specific Nii genes and transcriptional silencing of transgenes driven by the 19S or 35S promoter irrespective of their coding sequence and of their location in the genome. In this paper we show that silencing of transgenes carrying Nii1 sequences occurs irrespective of the promoter driving their expression and of their location within the genome. This phenomenon occurs in roots as well as in leaves although root Nii genes share only 84% identity with leaf-specific Nii1 sequences carried by the 271 locus. Conversely, transgenes carrying the bean Nii gene (which shares 76% identity with the tobacco Nii1 gene) escape silencing by the 271 locus. We also show that transgenes driven by the figwort mosaic virus 34S promoter (which shares 63% identity with the 35S promoter) also escape silencing by the 271 locus. Taken together, these results indicate that a high degree of sequence similarity is required between the sequences of the silencing locus and of the target (trans)genes for both transcriptional and post-transcriptional silencing.

  17. Using transgenic Caenorhabditis elegans in soil toxicity testing.

    PubMed

    Graves, Amber L; Boyd, Windy A; Williams, Phillip L

    2005-05-01

    Soil bioassays are important tools for evaluating toxicological effects within the terrestrial environment. The American Society for Testing and Materials E2172-01 Standard Guide outlines a method for conducting laboratory soil toxicity tests using the nematode Caenorhabditis elegans. This method is an efficient tool for extracting C. elegans from soil samples and can be carried out after a 24-h exposure period using relatively small amounts of soil. Drawbacks of this method include problems with (1) recovery of nematodes from soils containing a high percentage of organic matter, and (2) distinguishing indigenous nematode species from nematodes added for the laboratory test. Due in part to these issues, C. elegans has not been extensively accepted for use in soil testing. To address these concerns and improve upon the American Society for Testing and Materials method, this project focused on using transgenic strains of C. elegans carrying a GFP-expressing element. Lethality and behavior tests revealed that the transgenic nematodes respond similarly to the wild-type N2 strain, indicating that they can be used in the same manner in soil testing. The GFP marker is easily identifiable not only within soils containing a large amount of organic matter, but also in field-collected soils containing indigenous nematodes. These results support the use of transgenic GFP C. elegans in soil bioassays as a tool to further the reliability of laboratory toxicity tests.

  18. Differential regulation of laminin b1 transgene expression in the neonatal and adult mouse brain.

    PubMed

    Sharif, K A; Baker, H; Gudas, L J

    2004-01-01

    Laminins are the major glycoproteins present in basement membrane, a type of extracellular matrix. We showed that the LAMB1 gene, which encodes the laminin beta1 subunit, is transcriptionally activated by retinoic acid in embryonic stem cells. However, little information is available concerning LAMB1 developmental regulation and spatial expression in the adult mouse brain. In this study we used transgenic mice expressing different lengths of LAMB1 promoter driving beta-galactosidase to investigate developmental and adult transcriptional regulation in the regions of the brain in which the laminin beta1 protein is expressed. CNS expression was not observed in transgenic mice carrying a 1.4LAMB1betagal construct. Mice carrying a 2.5LAMB1betagal construct expressed the LAMB1 transgene, as assayed by X-gal staining, only in the molecular layer of the neonatal cerebellum. In contrast, a 3.9LAMB1betagal transgene showed broad regional expression in the adult mouse brain, including the hippocampus, entorhinal cortex, colliculi, striatum, and substantia nigra. Similar expression patterns were observed for the endogenous laminin beta1 protein and for the 3.9LAMB1betagal transgene, analyzed with an antibody against the beta-galactosidase protein. The 3.9LAMB1betagal transgene expression in the hippocampal tri-synaptic circuit suggests a role for the LAMB1 gene in learning and memory.

  19. Transgenic Arabidopsis Gene Expression System

    NASA Technical Reports Server (NTRS)

    Ferl, Robert; Paul, Anna-Lisa

    2009-01-01

    The Transgenic Arabidopsis Gene Expression System (TAGES) investigation is one in a pair of investigations that use the Advanced Biological Research System (ABRS) facility. TAGES uses Arabidopsis thaliana, thale cress, with sensor promoter-reporter gene constructs that render the plants as biomonitors (an organism used to determine the quality of the surrounding environment) of their environment using real-time nondestructive Green Fluorescent Protein (GFP) imagery and traditional postflight analyses.

  20. Transgene expression systems in the Triticeae cereals.

    PubMed

    Hensel, Götz; Himmelbach, Axel; Chen, Wanxin; Douchkov, Dimitar K; Kumlehn, Jochen

    2011-01-01

    The control of transgene expression is vital both for the elucidation of gene function and for the engineering of transgenic crops. Given the dominance of the Triticeae cereals in the agricultural economy of the temperate world, the development of well-performing transgene expression systems of known functionality is of primary importance. Transgenes can be expressed either transiently or stably. Transient expression systems based on direct or virus-mediated gene transfer are particularly useful in situations where the need is to rapidly screen large numbers of genes. However, an unequivocal understanding of gene function generally requires that a transgene functions throughout the plant's life and is transmitted through the sexual cycle, since this alone allows its effect to be decoupled from the plant's response to the generally stressful gene transfer event. Temporal, spatial and quantitative control of a transgene's expression depends on its regulatory environment, which includes both its promoter and certain associated untranslated region sequences. While many transgenic approaches aim to manipulate plant phenotype via ectopic gene expression, a transgene sequence can be also configured to down-regulate the expression of its endogenous counterpart, a strategy which exploits the natural gene silencing machinery of plants. In this review, current technical opportunities for controlling transgene expression in the Triticeae species are described. Apart from protocols for transient and stable gene transfer, the choice of promoters and other untranslated regulatory elements, we also consider signal peptides, as they too govern the abundance and particularly the sub-cellular localization of transgene products.

  1. Gene editing activity on extrachromosomal arrays in C. elegans transgenics.

    PubMed

    Falgowski, Kerry A; Kmiec, Eric B

    2011-04-15

    Gene editing by modified single-stranded oligonucleotides is a strategy aimed at inducing single base changes into the genome, generating a permanent genetic change. The work presented here explores gene editing capabilities in the model organism Caenorhabditis elegans. Current approaches to gene mutagenesis in C. elegans have been plagued by non-specificity and thus the ability to induce precise, directed alterations within the genome of C. elegans would offer a platform upon which structure/function analyses can be carried out. As such, several in vivo assay systems were developed to evaluate gene editing capabilities in C. elegans. Fluorescence was chosen as the selectable endpoint as fluorescence can be easily detected through the transparent worm body even from minimal expression. Two tissue specific fluorescent expression vectors containing either a GFP or mCherry transgene were mutagenized to create a single nonsense mutation within the open reading frame of each respective fluorescent gene. These served as the target site to evaluate the frequency of gene editing on extrachromosomal array transgenic lines. Extrachromosomal arrays can carry hundreds of copies of the transgene, therefore low frequency events (like those in the gene editing reaction) may be detected. Delivery of the oligonucleotide was accomplished by microinjection into the gonads of young adult worms in an effort to induce repair of the mutated fluorescent gene in the F1 progeny. Despite many microinjections on the transgenic strains with varying concentrations of ODNs, no gene editing events were detected. This result is consistent with the previous research, demonstrating the difficulties encountered in targeting embryonic stem cells and the pronuclei of single-celled embryos.

  2. Optimization of Biofuel Production From Transgenic Microalgae

    DTIC Science & Technology

    2013-02-27

    AFRL-OSR-VA-TR-2013-0145 OPTIMIZATION OF BIOFUEL PRODUCTION FROM TRANSGENIC MICROALGAE Richard Sayre Donald Danforth...Technical 20080815 to 20120630 OPTIMIZATION OF BIOFUEL PRODUCTION FROM TRANSGENIC MICROALGAE FA9550-08-1-0451 Richard Sayre Donald Danforth Plant...BIOFUEL PRODUCTION FROM TRANSGENIC MICROALGAE Grant/Contract Number: FA9550-08-1-0451 Reporting Period: Final Report Abstract: We have compared the

  3. Transfer of stem cells carrying engineered chromosomes with XY clone laser system.

    PubMed

    Sinko, Ildiko; Katona, Robert L

    2011-01-01

    Current transgenic technologies for gene transfer into the germline of mammals cause a random integration of exogenous naked DNA into the host genome that can generate undesirable position effects as well as insertional mutations. The vectors used to generate transgenic animals are limited by the amount of foreign DNA they can carry. Mammalian artificial chromosomes have large DNA-carrying capacity and ability to replicate in parallel with, but without integration into, the host genome. Hence they are attractive vectors for transgenesis, cellular protein production, and gene therapy applications as well. ES cells mediated chromosome transfer by conventional blastocyst injection has a limitation in unpredictable germline transmission. The demonstrated protocol of laser-assisted microinjection of artificial chromosome containing ES cells into eight-cell mouse embryos protocol described here can solve the problem for faster production of germline transchromosomic mice.

  4. A Brief Analysis of Sister Carrie's Character

    ERIC Educational Resources Information Center

    Yu, Hanying

    2010-01-01

    Carrie is always dreaming while the rocking chair is rocking again and again, this is the deep impression on us after we read "Sister Carrie" which is the first novel of Theodore Dreiser. In this novel the protagonist Sister Carrie is a controversial person. This paper tries to analyze the character of Sister Carrie in order to find out…

  5. Xenopus transgenics: methods using transposons.

    PubMed

    Kelley, Clair M; Yergeau, Donald A; Zhu, Haiqing; Kuliyev, Emin; Mead, Paul E

    2012-01-01

    The generation of transgenic animals is an essential tool for many genetic strategies. DNA "cut-and-paste" transposon systems can be used to efficiently modify the Xenopus genome. The DNA transposon substrate, harbored on a circularized plasmid, is co-injected into fertilized Xenopus embryos at the one-cell stage together with mRNA encoding the cognate transposase enzyme. The cellular machinery rapidly translates the exogenous mRNA to produce active transposase enzyme that catalyzes excision of the transposon substrate from the plasmid and stable integration into the genomic DNA.

  6. Virus-derived transgenes expressing hairpin RNA give immunity to Tobacco mosaic virus and Cucumber mosaic virus

    PubMed Central

    2011-01-01

    Background An effective method for obtaining resistant transgenic plants is to induce RNA silencing by expressing virus-derived dsRNA in plants and this method has been successfully implemented for the generation of different plant lines resistant to many plant viruses. Results Inverted repeats of the partial Tobacco mosaic virus (TMV) movement protein (MP) gene and the partial Cucumber mosaic virus (CMV) replication protein (Rep) gene were introduced into the plant expression vector and the recombinant plasmids were transformed into Agrobacterium tumefaciens. Agrobacterium-mediated transformation was carried out and three transgenic tobacco lines (MP16-17-3, MP16-17-29 and MP16-17-58) immune to TMV infection and three transgenic tobacco lines (Rep15-1-1, Rep15-1-7 and Rep15-1-32) immune to CMV infection were obtained. Virus inoculation assays showed that the resistance of these transgenic plants could inherit and keep stable in T4 progeny. The low temperature (15℃) did not influence the resistance of transgenic plants. There was no significant correlation between the resistance and the copy number of the transgene. CMV infection could not break the resistance to TMV in the transgenic tobacco plants expressing TMV hairpin MP RNA. Conclusions We have demonstrated that transgenic tobacco plants expressed partial TMV movement gene and partial CMV replicase gene in the form of an intermolecular intron-hairpin RNA exhibited complete resistance to TMV or CMV infection. PMID:21269519

  7. Cutaneous lymphoproliferation and lymphomas in interleukin 7 transgenic mice

    PubMed Central

    1993-01-01

    To investigate the role of interleukin 7 (IL-7) in the development of the lymphoid system, we have generated two lines of transgenic mice carrying an IL-7 cDNA fused to an immunoglobulin heavy chain promoter and enhancer. This transgene is expressed in the bone marrow, lymph nodes, spleen, thymus, and skin provoking a perturbation of T cell development characterized by a marked reduction of CD4+ CD8+ (double- positive) thymocytes. Quite unexpectedly, however, both lines also develop a progressive cutaneous disorder involving a dermal lymphoid infiltrate that results in progressive alopecia, hyperkeratosis, and exfoliation. Although the infiltrate is primarily composed of T lineage cells, its development is not impeded in the athymic nu/nu background. Furthermore, the phenotype can be transmitted horizontally by transplanting lymphoid tissues or skin to syngeneic wild-type mice. Thus, the phenotype is conveyed by skin-homing, mobile cells (presumably the infiltrating lymphocytes) in a cell-autonomous fashion. In addition to the skin phenotype, this transgene also provokes the development of a lymphoproliferative disorder that induces B and T cell lymphomas within the first 4 mo of life. These findings suggest potential physiologic actions of IL-7 in T cell development and in cutaneous immunity. They also demonstrate that IL-7 can act as an oncogene in the living organism. PMID:7678850

  8. Molecular Characterization of Transgenic Events Using Next Generation Sequencing Approach

    PubMed Central

    Mammadov, Jafar; Ye, Liang; Soe, Khaing; Richey, Kimberly; Cruse, James; Zhuang, Meibao; Gao, Zhifang; Evans, Clive; Rounsley, Steve; Kumpatla, Siva P.

    2016-01-01

    Demand for the commercial use of genetically modified (GM) crops has been increasing in light of the projected growth of world population to nine billion by 2050. A prerequisite of paramount importance for regulatory submissions is the rigorous safety assessment of GM crops. One of the components of safety assessment is molecular characterization at DNA level which helps to determine the copy number, integrity and stability of a transgene; characterize the integration site within a host genome; and confirm the absence of vector DNA. Historically, molecular characterization has been carried out using Southern blot analysis coupled with Sanger sequencing. While this is a robust approach to characterize the transgenic crops, it is both time- and resource-consuming. The emergence of next-generation sequencing (NGS) technologies has provided highly sensitive and cost- and labor-effective alternative for molecular characterization compared to traditional Southern blot analysis. Herein, we have demonstrated the successful application of both whole genome sequencing and target capture sequencing approaches for the characterization of single and stacked transgenic events and compared the results and inferences with traditional method with respect to key criteria required for regulatory submissions. PMID:26908260

  9. Gene flow from transgenic to nontransgenic soybean plants in the Cerrado region of Brazil.

    PubMed

    Abud, S; de Souza, P I M; Vianna, G R; Leonardecz, E; Moreira, C T; Faleiro, F G; Júnior, J N; Monteiro, P M F O; Rech, E L; Aragão, F J L

    2007-06-30

    Evaluation of transgenic crops under field conditions is a fundamental step for the production of genetically engineered varieties. In order to determine if there is pollen dispersal from transgenic to nontransgenic soybean plants, a field release experiment was conducted in the Cerrado region of Brazil. Nontransgenic plants were cultivated in plots surrounding Roundup Ready transgenic plants carrying the cp4 epsps gene, which confers herbicide tolerance against glyphosate herbicide, and pollen dispersal was evaluated by checking for the dominant gene. The percentage of cross-pollination was calculated as a fraction of herbicide-tolerant and -nontolerant plants. The greatest amount of transgenic pollen dispersion was observed in the first row, located at one meter from the central (transgenic) plot, with a 0.52% average frequency. The frequency of pollen dispersion decreased to 0.12% in row 2, reaching 0% when the plants were up to 10 m distance from the central plot. Under these conditions pollen flow was higher for a short distance. This fact suggests that the management necessary to avoid cross-pollination from transgenic to nontransgenic plants in the seed production fields should be similar to the procedures currently utilized to produce commercial seeds.

  10. Transgenic mouse model of hemifacial microsomia: Cloning and characterization of insertional mutation region on chromosome 10

    SciTech Connect

    Naora, Hiroyuki; Otani, Hiroki; Tanaka, Osamu

    1994-10-01

    The 643 transgenic mouse line carries an autosomal dominant insertional mutation that results in hemifacial microsomia (HFM), including microtia and/or abnormal biting. In this paper, we characterize the transgene integration site in transgenic mice and preintegration site of wildtype mice. The locus, designated Hfm (hemifacial microsomia-associated locus), was mapped to chromosome 10, B1-3, by chromosome in situ hybridization. We cloned the transgene insertion site from the transgenic DNA library. By using the 5{prime} and 3{prime} flanking sequences, the preintegration region was isolated. The analysis of these regions showed that a deletion of at least 23 kb DNA occurred in association with the transgene integration. Evolutionarily conserved regions were detected within and beside the deleted region. The result of mating between hemizygotes suggests that the phenotype of the homozygote is lethality in the prenatal period. These results suggests that the Hfm locus is necessary for prenatal development and that this strain is a useful animal model for investigating the genetic predisposition to HFM in humans.

  11. Tol2-Mediated Generation of a Transgenic Haplochromine Cichlid, Astatotilapia burtoni

    PubMed Central

    Fernald, Russell D.

    2013-01-01

    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. PMID:24204902

  12. Nematode neuropeptides as transgenic nematicides.

    PubMed

    Warnock, Neil D; Wilson, Leonie; Patten, Cheryl; Fleming, Colin C; Maule, Aaron G; Dalzell, Johnathan J

    2017-02-01

    Plant parasitic nematodes (PPNs) seriously threaten global food security. Conventionally an integrated approach to PPN management has relied heavily on carbamate, organophosphate and fumigant nematicides which are now being withdrawn over environmental health and safety concerns. This progressive withdrawal has left a significant shortcoming in our ability to manage these economically important parasites, and highlights the need for novel and robust control methods. Nematodes can assimilate exogenous peptides through retrograde transport along the chemosensory amphid neurons. Peptides can accumulate within cells of the central nerve ring and can elicit physiological effects when released to interact with receptors on adjoining cells. We have profiled bioactive neuropeptides from the neuropeptide-like protein (NLP) family of PPNs as novel nematicides, and have identified numerous discrete NLPs that negatively impact chemosensation, host invasion and stylet thrusting of the root knot nematode Meloidogyne incognita and the potato cyst nematode Globodera pallida. Transgenic secretion of these peptides from the rhizobacterium, Bacillus subtilis, and the terrestrial microalgae Chlamydomonas reinhardtii reduce tomato infection levels by up to 90% when compared with controls. These data pave the way for the exploitation of nematode neuropeptides as a novel class of plant protective nematicide, using novel non-food transgenic delivery systems which could be deployed on farmer-preferred cultivars.

  13. Nematode neuropeptides as transgenic nematicides

    PubMed Central

    Patten, Cheryl; Fleming, Colin C.; Maule, Aaron G.

    2017-01-01

    Plant parasitic nematodes (PPNs) seriously threaten global food security. Conventionally an integrated approach to PPN management has relied heavily on carbamate, organophosphate and fumigant nematicides which are now being withdrawn over environmental health and safety concerns. This progressive withdrawal has left a significant shortcoming in our ability to manage these economically important parasites, and highlights the need for novel and robust control methods. Nematodes can assimilate exogenous peptides through retrograde transport along the chemosensory amphid neurons. Peptides can accumulate within cells of the central nerve ring and can elicit physiological effects when released to interact with receptors on adjoining cells. We have profiled bioactive neuropeptides from the neuropeptide-like protein (NLP) family of PPNs as novel nematicides, and have identified numerous discrete NLPs that negatively impact chemosensation, host invasion and stylet thrusting of the root knot nematode Meloidogyne incognita and the potato cyst nematode Globodera pallida. Transgenic secretion of these peptides from the rhizobacterium, Bacillus subtilis, and the terrestrial microalgae Chlamydomonas reinhardtii reduce tomato infection levels by up to 90% when compared with controls. These data pave the way for the exploitation of nematode neuropeptides as a novel class of plant protective nematicide, using novel non-food transgenic delivery systems which could be deployed on farmer-preferred cultivars. PMID:28241060

  14. Perspective on the combined use of an independent transgenic sexing and a multifactorial reproductive sterility system to avoid resistance development against transgenic Sterile Insect Technique approaches

    PubMed Central

    2014-01-01

    Background The Sterile Insect Technique (SIT) is an accepted species-specific genetic control approach that acts as an insect birth control measure, which can be improved by biotechnological engineering to facilitate its use and widen its applicability. First transgenic insects carrying a single killing system have already been released in small scale trials. However, to evade resistance development to such transgenic approaches, completely independent ways of transgenic killing should be established and combined. Perspective Most established transgenic sexing and reproductive sterility systems are based on the binary tTA expression system that can be suppressed by adding tetracycline to the food. However, to create 'redundant killing' an additional independent conditional expression system is required. Here we present a perspective on the use of a second food-controllable binary expression system - the inducible Q system - that could be used in combination with site-specific recombinases to generate independent transgenic killing systems. We propose the combination of an already established transgenic embryonic sexing system to meet the SIT requirement of male-only releases based on the repressible tTA system together with a redundant male-specific reproductive sterility system, which is activated by Q-system controlled site-specific recombination and is based on a spermatogenesis-specifically expressed endonuclease acting on several species-specific target sites leading to chromosome shredding. Conclusion A combination of a completely independent transgenic sexing and a redundant reproductive male sterility system, which do not share any active components and mediate the induced lethality by completely independent processes, would meet the 'redundant killing' criteria for suppression of resistance development and could therefore be employed in large scale long-term suppression programs using biotechnologically enhanced SIT. PMID:25471733

  15. Transgenic Biofuel Feedstocks and Strategies for Biocontainment

    Technology Transfer Automated Retrieval System (TEKTRAN)

    There are several reasons to believe that transgenic plant feedstocks will be required to realize the full economic and environmental benefits of cellulosic and other biofuels. Much of the commercialization potential for the use of transgenic plant cellulosic feedstocks may be impacted by regulatio...

  16. An affinity-based genome walking method to find transgene integration loci in transgenic genome.

    PubMed

    Thirulogachandar, V; Pandey, Prachi; Vaishnavi, C S; Reddy, Malireddy K

    2011-09-15

    Identifying a good transgenic event from the pool of putative transgenics is crucial for further characterization. In transgenic plants, the transgene can integrate in either single or multiple locations by disrupting the endogenes and/or in heterochromatin regions causing the positional effect. Apart from this, to protect the unauthorized use of transgenic plants, the signature of transgene integration for every commercial transgenic event needs to be characterized. Here we show an affinity-based genome walking method, named locus-finding (LF) PCR (polymerase chain reaction), to determine the transgene flanking sequences of rice plants transformed by Agrobacterium tumefaciens. LF PCR includes a primary PCR by a degenerated primer and transfer DNA (T-DNA)-specific primer, a nested PCR, and a method of enriching the desired amplicons by using a biotin-tagged primer that is complementary to the T-DNA. This enrichment technique separates the single strands of desired amplicons from the off-target amplicons, reducing the template complexity by several orders of magnitude. We analyzed eight transgenic rice plants and found the transgene integration loci in three different chromosomes. The characteristic illegitimate recombination of the Agrobacterium sp. was also observed from the sequenced integration loci. We believe that the LF PCR should be an indispensable technique in transgenic analysis.

  17. Field Performance of Transgenic Sugarcane Lines Resistant to Sugarcane Mosaic Virus

    PubMed Central

    Yao, Wei; Ruan, Miaohong; Qin, Lifang; Yang, Chuanyu; Chen, Rukai; Chen, Baoshan; Zhang, Muqing

    2017-01-01

    Sugarcane mosaic disease is mainly caused by the sugarcane mosaic virus (SCMV), which can significantly reduce stalk yield and sucrose content of sugarcane in the field. Coat protein mediated protection (CPMP) is an effective strategy to improve virus resistance. A 2-year field study was conducted to compare five independent transgenic sugarcane lines carrying the SCMV-CP gene (i.e., B2, B36, B38, B48, and B51) with the wild-type parental clone Badila (WT). Agronomic performance, resistance to SCMV infection, and transgene stability were evaluated and compared with the wild-type parental clone Badila (WT) at four experimental locations in China across two successive seasons, i.e., plant cane (PC) and 1st ratoon cane (1R). All transgenic lines derived from Badila had significantly greater tons of cane per hectare (TCH) and tons of sucrose per hectare (TSH) as well as lower SCMV disease incidence than those from Badila in the PC and 1R crops. The transgenic line B48 was highly resistant to SCMV with less than 3% incidence of infection. The recovery phenotype of transgenic line B36 was infected soon after virus inoculation, but the subsequent leaves showed no symptoms of infection. Most control plants developed symptoms that persisted and spread throughout the plant with more than 50% incidence. B48 recorded an average of 102.72 t/ha, which was 67.2% more than that for Badila. The expression of the transgene was stable over many generations with vegetative propagation. These results show that SCMV-resistant transgenic lines derived from Badila can provide resistant germplasm for sugarcane breeding and can also be used to study virus resistance mechanisms. This is the first report on the development and field performance of transgenic sugarcane plants that are resistant to SCMV infection in China. PMID:28228765

  18. Accumulation of nickel in transgenic tobacco

    NASA Astrophysics Data System (ADS)

    Sidik, Nik Marzuki; Othman, Noor Farhan

    2013-11-01

    The accumulation of heavy metal Ni in the roots and leaves of four T1 transgenic lines of tobacco (T(1)20E, T(1)24C, T(1)18B1 and T(1)20B) expressing eiMT1 from E.indica was assessed. The aim of the study was to investigate the level of Ni accumulation in the leaves and roots of each transgenic lines and to evaluate the eligibility of the plants to be classified as a phytoremediation agent. All of the transgenic lines showed different ability in accumulating different metals and has translocation factor (TF) less than 1 (TF<1) at all levels of metal treatment. Among the 4 transgenic lines, transgenic line T(1)24C showed the highest accumulation of Ni (251.9 ± 0.014 mg/kg) and the lowest TF value (TFT(1)24C=0.0875) at 60 ppm Ni.

  19. A Transgenic Durum Wheat Line that is Free of Marker Genes and Expresses 1dy10

    Technology Transfer Automated Retrieval System (TEKTRAN)

    We used a combination of “clean gene” technology and positive selection to generate transgenic durum wheat lines free of herbicide and antibiotic resistance marker genes. Biolistic transformation experiments were carried out using three “minimal gene cassettes” consisting of linear DNA fragments exc...

  20. Genomic evaluation of oxalate-degrading transgenic soybean in response to Sclerotinia sclerotiorum infection

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Oxalate oxidases catalyze the degradation of oxalic acid (OA). Highly resistant transgenic soybean carrying an oxalate oxidase (OxO) gene and its susceptible parent soybean line, AC Colibri, were tested for genome-wide gene expression in response to the necrotrophic, OA producing pathogen Sclerotini...

  1. Efficient Production of Fluorescent Transgenic Rats using the piggyBac Transposon

    PubMed Central

    Li, Tianda; Shuai, Ling; Mao, Junjie; Wang, Xuepeng; Wang, Mei; Zhang, Xinxin; Wang, Leyun; Li, Yanni; Li, Wei; Zhou, Qi

    2016-01-01

    Rats with fluorescent markers are of great value for studies that trace lineage-specific development, particularly those assessing the differentiation potential of embryonic stem cells (ESCs). The piggyBac (PB) transposon is widely used for the efficient introduction of genetic modifications into genomes, and has already been successfully used to produce transgenic mice and rats. Here, we generated transgenic rats carrying either the desRed fluorescent protein (RFP) gene or the enhanced green fluorescent protein (eGFP) gene by injecting pronuclei with PB plasmids. We showed that the transgenic rats expressed the RFP or eGFP gene in many organs and had the capability to transmit the marker gene to the next generation through germline integration. In addition, rat embryonic stem cells (ESCs) carrying an RFP reporter gene can be derived from the blastocysts of the transgenic rats. Moreover, the RFP gene can be detected in chimeras derived from RFP ESCs via blastocyst injection. This work suggests that PB-mediated transgenesis is a powerful tool to generate transgenic rats expressing fluorescent proteins with high efficiency, and this technique can be used to derive rat ESCs expressing a reporter protein. PMID:27624004

  2. The beta 1 integrin distal promoter is developmentally regulated in transgenic mice.

    PubMed

    Hirsch, E; Balzac, F; Pastore, C; Tarone, G; Silengo, L; Altruda, F

    1993-12-01

    Transgenic mice harbouring 5' flanking sequences of the human beta 1 integrin gene linked to the Escherichia coli lacZ gene have been generated to examine spatial and temporal distribution of the promoter activity during development. Our previous data showed that this regulatory region is composed by two promoters, called distal and proximal, located closely on the human genome. To determine the role of each promoter region during development we generated transgenic mice using these two sequences linked to the lacZ reporter gene. Their analysis shows that these two sequences, as determined by in vitro studies, have different efficiencies in promoting transcription. Actually mice carrying the proximal promoter region exhibit a weak lacZ expression resulting in an undetectable beta-galactosidase activity in both embryonic and adult tissues. On the other hand, transgenic mice carrying the distal promoter express beta-galactosidase at high efficiency during embryonic development. The pattern of transgene expression is consistent with the localization of beta 1 protein on mouse embryos evidenced by immunohistochemistry. Moreover the distal promoter is subjected to a temporal modulation since in adult transgenic mice lacZ expression decreases to a level detected only by RT-PCR analysis. We have determined a similar down-regulation analysing by Northern blot beta 1 mRNA in adult and embryonic organs such as heart and gut.

  3. Taking on Titan: Meet Carrie Anderson

    NASA Video Gallery

    When she was a little girl, Carrie Anderson dreamed of becoming an astronomer. Now, as a space scientist at NASA Goddard Space Flight Center, Carrie studies the atmosphere on Titan: one of Saturn's...

  4. Production of transgenic canine embryos using interspecies somatic cell nuclear transfer.

    PubMed

    Hong, So Gun; Oh, Hyun Ju; Park, Jung Eun; Kim, Min Jung; Kim, Geon A; Koo, Ok Jae; Jang, Goo; Lee, Byeong Chun

    2012-02-01

    Somatic cell nuclear transfer (SCNT) has emerged as an important tool for producing transgenic animals and deriving transgenic embryonic stem cells. The process of SCNT involves fusion of in vitro matured oocytes with somatic cells to make embryos that are transgenic when the nuclear donor somatic cells carry 'foreign' DNA and are clones when all the donor cells are genetically identical. However, in canines, it is difficult to obtain enough mature oocytes for successful SCNT due to the very low efficiency of in vitro oocyte maturation in this species that hinders canine transgenic cloning. One solution is to use oocytes from a different species or even a different genus, such as bovine oocytes, that can be matured easily in vitro. Accordingly, the aim of this study was: (1) to establish a canine fetal fibroblast line transfected with the green fluorescent protein (GFP) gene; and (2) to investigate in vitro embryonic development of canine cloned embryos derived from transgenic and non-transgenic cell lines using bovine in vitro matured oocytes. Canine fetal fibroblasts were transfected with constructs containing the GFP and puromycin resistance genes using FuGENE 6®. Viability levels of these cells were determined by the MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide] assay. Interspecies SCNT (iSCNT) embryos from normal or transfected cells were produced and cultured in vitro. The MTT measurement of GFP-transfected fetal fibroblasts (mean OD = 0.25) was not significantly different from non-transfected fetal fibroblasts (mean OD = 0.35). There was no difference between transgenic iSCNT versus non-transgenic iSCNT embryos in terms of fusion rates (73.1% and 75.7%, respectively), cleavage rates (69.7% vs. 73.8%) and development to the 8-16-cell stage (40.1% vs. 42.7%). Embryos derived from the transfected cells completely expressed GFP at the 2-cell, 4-cell, and 8-16-cell stages without mosaicism. In summary, our results demonstrated that

  5. 25 CFR 167.6 - Carrying capacities.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 25 Indians 1 2011-04-01 2011-04-01 false Carrying capacities. 167.6 Section 167.6 Indians BUREAU... Carrying capacities. (a) The Commissioner of Indian Affairs on June 26, 1943, promulgated the authorized carrying capacity for each land management district of the Navajo Reservation. (b) Recommended...

  6. 25 CFR 167.6 - Carrying capacities.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 25 Indians 1 2012-04-01 2011-04-01 true Carrying capacities. 167.6 Section 167.6 Indians BUREAU OF... capacities. (a) The Commissioner of Indian Affairs on June 26, 1943, promulgated the authorized carrying... carrying capacities shall be referred by the Superintendent to District Grazing Committee, Central...

  7. 25 CFR 167.6 - Carrying capacities.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 25 Indians 1 2013-04-01 2013-04-01 false Carrying capacities. 167.6 Section 167.6 Indians BUREAU... Carrying capacities. (a) The Commissioner of Indian Affairs on June 26, 1943, promulgated the authorized carrying capacity for each land management district of the Navajo Reservation. (b) Recommended...

  8. 25 CFR 167.6 - Carrying capacities.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 25 Indians 1 2010-04-01 2010-04-01 false Carrying capacities. 167.6 Section 167.6 Indians BUREAU... Carrying capacities. (a) The Commissioner of Indian Affairs on June 26, 1943, promulgated the authorized carrying capacity for each land management district of the Navajo Reservation. (b) Recommended...

  9. [Research progress on environmental carrying capacity].

    PubMed

    Wang, Jian; Sun, Tieheng; Li, Peijun; Li, Fayun

    2005-04-01

    To study the theories and quantification methods of environmental carrying capacity is of significance in reality for directing human beings economic behaviors and harmonizing the relationships between social development and environment. In this paper, the definition of environmental carrying capacity was introduced from the aspects of "capacity", "threshold" and "capability", with the main characteristics of objective and subjective, regional and temporal, and dynamic and adjustable, and its research progress was reviewed. On the basis of these, the quantification methods of environmental carrying capacity, including exponential assessment, carrying rate assessment, system dynamics, and multi-objective optimization, were analyzed, and the research perspectives of environmental carrying capacity were discussed.

  10. Promoter methylation and histone modifications affect the expression of the exogenous DsRed gene in transgenic goats.

    PubMed

    Nuo, M T; Yuan, J L; Yang, W L; Gao, X Y; He, N; Liang, H; Cang, M; Liu, D J

    2016-08-29

    Transgene silencing, which is common in transgenic plants and animals, limits the generation and application of genetically modified organisms, and is associated with the exogenous gene copy number, the methylation status of its promoters, and histone modification abnormalities. Here, we analyzed the expression of the exogenous gene DsRed and the methylation status of its cytomegalovirus (CMV) promoter in six healthy transgenic cashmere goats and transgenic nuclear donor cells. The CMV promoter exhibited high methylation levels (74.4-88.2%) in four of the goats, a moderate methylation level (58.7%) in one, and a low methylation level (21.2%) in one, while the methylation level of the transgenic nuclear donor cells was comparatively low (14.3%). DsRed expression was negatively correlated with promoter methylation status. Transgenic cashmere goats carried one to three copies of the CMV promoter fragment and one to six copies of the DsRed fragment, but copy number showed no obvious correlation with DsRed expression. After treatment with the methylation inhibitor 5-azacytidine, DsRed expression in transgenic goat cells significantly increased and CMV promoter methylation significantly decreased; this indicated an inverse correlation between promoter methylation status and DsRed expression. After treatment with the histone deacetylase inhibitor trichostatin A, DsRed expression increased, indicating that an abnormal histone modification in transgenic goats is also involved in exogenous gene silencing. These findings indicate the potential of trichostatin A and 5-azacytidine to rescue the biological activity of silenced exogenous transgenes in adult-derived transgenic cells under culture conditions.

  11. Risk assessment of transgenic apomictic tetraploid bahiagrass, cytogenetics, breeding behavior and performance of intra-specific hybrids.

    PubMed

    Sandhu, Sukhpreet; James, Victoria A; Quesenberry, Kenneth H; Altpeter, Fredy

    2009-11-01

    Pollen-mediated gene transfer from stress tolerant or herbicide-resistant transgenic plants may cause environmental or agronomic problems. Apomictic seed production found in some bahiagrass cultivars may serve as a natural transgene containment system. Under greenhouse conditions, the average gene transfer frequency from an herbicide-resistant apomictic tetraploid to a population of sexual diploid bahiagrass genotypes or apomictic tetraploid bahiagrass was 0.16% when the transgenic pollen donor was placed at 0.5-1.5 m distance from the non-transgenic pollen receptors. The herbicide-resistant hybrids were characterized for transgene integration, expression and ploidy, by Southern blot analysis, immuno-chromatography and flow cytometry, respectively. Hybrids resulting from open pollination of non-transgenic diploid female plants with transgenic tetraploid male plants were triploids or near-triploids, with 2n = 26-34. These hybrids displayed a wide range of phenotypic variability, including some non-persistent or non-flowering dwarf-type hybrids with good vigor, or hybrids with vegetative growth similar to non-transgenic plants, but with significantly reduced seed set. Non-flowering aneu-triploids with good vigor/field performance will provide the highest level of transgene containment. Embryo sac analysis of pollinated spikelets confirmed a high proportion of aborted ovules. An apospory-linked RFLP marker was detected in 13 of the 15 near-triploid hybrids. All flowering aneuploid hybrids displayed significantly reduced seed set, and none of the sexual near-triploid hybrids produced any seeds. All tetraploid gene transfer events carried the apospory-linked RFLP marker, suggesting that despite the presence of the aposporus locus, a low degree of sexuality co-exists in apomictic tetraploid cultivars. Thus, tetraploid apomictic bahiagrass does not provide complete transgene containment, although intra-specific gene transfer is drastically reduced compared to sexually

  12. Development of Cotton leaf curl virus resistant transgenic cotton using antisense ßC1 gene.

    PubMed

    Sohrab, Sayed Sartaj; Kamal, Mohammad A; Ilah, Abdul; Husen, Azamal; Bhattacharya, P S; Rana, D

    2016-05-01

    Cotton leaf curl virus (CLCuV) is a serious pathogen causing leaf curl disease and affecting the cotton production in major growing areas. The transgenic cotton (Gossypium hirsutum cv. Coker 310) plants were developed by using βC1 gene in antisense orientation gene driven by Cauliflower mosaic virus-35S promoter and nos (nopaline synthase) terminator and mediated by Agrobacterium tumefaciens transformation and somatic embryogenesis system. Molecular confirmation of the transformants was carried out by polymerase chain reaction (PCR) and Southern blot hybridization. The developed transgenic and inoculated plants remained symptomless till their growth period. In conclusion, the plants were observed as resistant to CLCuV.

  13. Glyphostate-drift but not herbivory alters the rate of transgene flow from single and stacked trait transgenic canola (Brassica napus L.) to non-transgenic B. napus and B. rapa

    EPA Science Inventory

    While transgenic plants can offer agricultural benefits, the escape of transgenes out of crop fields is a major environmental concern. Escape of transgenic herbicide resistance has occurred between transgenic Brassica napus (canola) and weedy species in numerous locations. In t...

  14. Expression of multiple proteins in transgenic plants

    DOEpatents

    Vierstra, Richard D.; Walker, Joseph M.

    2002-01-01

    A method is disclosed for the production of multiple proteins in transgenic plants. A DNA construct for introduction into plants includes a provision to express a fusion protein of two proteins of interest joined by a linking domain including plant ubiquitin. When the fusion protein is produced in the cells of a transgenic plant transformed with the DNA construction, native enzymes present in plant cells cleave the fusion protein to release both proteins of interest into the cells of the transgenic plant. Since the proteins are produced from the same fusion protein, the initial quantities of the proteins in the cells of the plant are approximately equal.

  15. Generation of red fluorescent protein transgenic dogs.

    PubMed

    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.

  16. Transgene integration and chromosome alterations in two transgenic lines of tritordeum.

    PubMed

    Barro, F; Martín, A; Cabrera, A

    2003-01-01

    Plants from two transgenic lines of tritordeum (an amphiploid between Triticum turgidum cv. durum and Hordeumn chilense) have been analyzed by fluorescence in-situ hybridization (FISH) to characterize the transgene integration sites and chromosome rearrangements. Transgenic lines were transformed in two different events with the genes encoding for the high-molecular-weight glutenin subunits (HMW-GS), 1Ax1 and/or 1Dx5. Three integration sites and four translocations were detected. All three integration sites were located on chromosome segments of Hordeum chilense translocated into wheat chromosomes. No translocations from wheat into H. chilense chromosomes were observed. Both HMW-GS transgenes were expressed at high levels in the endosperm of transgenic plants. The analysis by FISH of transgenic plants allowed the early detection of homozygous and heterozygous plants. The consequences and implications of translocations on breeding are discussed.

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

    PubMed

    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.

  18. Transgenic plants with enhanced growth characteristics

    DOEpatents

    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.

  19. AN APPROACH TO TRANSGENIC CROP MONITORING

    EPA Science Inventory

    Remote sensing by aerial or satellite images may provide a method of identifying transgenic pesticidal crop distribution in the landscape. Genetically engineered crops containing bacterial gene(s) that express an insecticidal protein from Bacillus thuringiensis (Bt) are regulated...

  20. Phenotyping transgenic wheat for drought resistance.

    PubMed

    Saint Pierre, Carolina; Crossa, José L; Bonnett, David; Yamaguchi-Shinozaki, Kazuko; Reynolds, Matthew P

    2012-03-01

    Realistic experimental protocols to screen for drought adaptation in controlled conditions are crucial if high throughput phenotyping is to be used for the identification of high performance lines, and is especially important in the evaluation of transgenes where stringent biosecurity measures restrict the frequency of open field trials. Transgenic DREB1A-wheat events were selected under greenhouse conditions by evaluating survival and recovery under severe drought (SURV) as well as for water use efficiency (WUE). Greenhouse experiments confirmed the advantages of transgenic events in recovery after severe water stress. Under field conditions, the group of transgenic lines did not generally outperform the controls in terms of grain yield under water deficit. However, the events selected for WUE were identified as lines that combine an acceptable yield-even higher yield (WUE-11) under well irrigated conditions-and stable performance across the different environments generated by the experimental treatments.

  1. Engineering the Caenorhabditis elegans genome by Mos1-induced transgene-instructed gene conversion.

    PubMed

    Robert, Valérie J P

    2012-01-01

    Mos1-induced transgene-instructed gene conversion (MosTIC) is a technique of choice to engineer the genome of the nematode Caenorhabditis elegans. MosTIC is initiated by the excision of Mos1, a DNA transposon of the Tc1/Mariner super family that can be mobilized in the germ line of C. elegans. Mos1 excision creates a DNA double-strand break that is repaired by several cellular mechanisms, including transgene-instructed gene conversion. For MosTIC, the transgenic repair template used by the gene conversion machinery is made of sequences that share DNA homologies with the genomic region to engineer and carries the modifications to be introduced in the genome. In this chapter, we present two MosTIC protocols routinely used.

  2. Health Considerations Regarding Horizontal Transfer of Microbial Transgenes Present in Genetically Modified Crops

    PubMed Central

    Kleter, Gijs A.

    2005-01-01

    The potential effects of horizontal gene transfer on human health are an important item in the safety assessment of genetically modified organisms. Horizontal gene transfer from genetically modified crops to gut microflora most likely occurs with transgenes of microbial origin. The characteristics of microbial transgenes other than antibiotic-resistance genes in market-approved genetically modified crops are reviewed. These characteristics include the microbial source, natural function, function in genetically modified crops, natural prevalence, geographical distribution, similarity to other microbial genes, known horizontal transfer activity, selective conditions and environments for horizontally transferred genes, and potential contribution to pathogenicity and virulence in humans and animals. The assessment of this set of data for each of the microbial genes reviewed does not give rise to health concerns. We recommend including the above-mentioned items into the premarket safety assessment of genetically modified crops carrying transgenes other than those reviewed in the present study. PMID:16489267

  3. Stress-induced synthesis of proline confers tolerance to water deficit in transgenic wheat.

    PubMed

    Vendruscolo, Eliane Cristina Gruszka; Schuster, Ivan; Pileggi, Marcos; Scapim, Carlos Alberto; Molinari, Hugo Bruno Correa; Marur, Celso Jamil; Vieira, Luiz Gonzaga Esteves

    2007-10-01

    Water deficit is one of the main abiotic factors that affect spring wheat planted in subtropical regions. Accumulation of proline appears to be a promising approach to maintain the productivity of plants under stress condition. However, morphological alterations and growth reduction are observed in transgenic plants carrying genes coding for osmoprotectants controlled by constitutive promoters. We report here the effects of water deficit on wheat plants transformed with the Vigna aconitifolia Delta(1)-pyrroline-5-carboxylate synthetase (P5CS) cDNA that encodes the key regulatory enzyme in proline biosynthesis, under the control of a stress-induced promoter complex-AIPC. Transgenic wheat plants submitted to 15 days of water shortage presented a distinct response. We have found that drought resulted in the accumulation of proline. The tolerance to water deficit observed in transgenic plants was mainly due to protection mechanisms against oxidative stress and not caused by osmotic adjustment.

  4. Safety assessment of meat from transgenic cattle by 90-day feeding study in rats.

    PubMed

    Liu, Shan; Li, Chen-Xi; Feng, Xiao-Lian; Wang, Hui-Ling; Liu, Hai-Bo; Zhi, Yuan; Geng, Gui-Ying; Zhao, Jie; Xu, Hai-Bin

    2013-07-01

    The study was carried out to evaluate the subchronic toxicity of meat derived from human lactoferrin gene-modified cattle in male and female Wistar rats. Rats were fed 5% or 10% transgenic meat diet, 5% or 10% conventional meat diet, or AIN93G diet for 90 days. During the study, body weight and food consumption were weighed weekly and clinical observations were conducted daily. At the end of the study, urinary examination, hematology and blood biochemistry examination, macroscopic and microscopic examinations were performed. There were no biologically significant differences in these factors between the rat groups fed transgenic meat diet and conventional meat diet. Therefore, the present 90-day rodent feeding study suggests that meat derived from the transgenic cattle is equivalent to meat from conventional cattle in use as dietary supplements.

  5. Assessing the Feasibility of Controlling Aedes aegypti with Transgenic Methods: A Model-Based Evaluation

    PubMed Central

    Legros, Mathieu; Xu, Chonggang; Okamoto, Kenichi; Scott, Thomas W.; Morrison, Amy C.; Lloyd, Alun L.; Gould, Fred

    2012-01-01

    Suppression of dengue and malaria through releases of genetically engineered mosquitoes might soon become feasible. Aedes aegypti mosquitoes carrying a conditionally lethal transgene have recently been used to suppress local vector populations in small-scale field releases. Prior to releases of transgenic insects on a wider scale, however, most regulatory authorities will require additional evidence that suppression will be effective in natural heterogeneous habitats. We use a spatially explicit stochastic model of an Ae. aegypti population in Iquitos, Peru, along with an uncertainty analysis of its predictions, to quantitatively assess the outcome of varied operational approaches for releases of transgenic strains with conditional death of females. We show that population elimination might be an unrealistic objective in heterogeneous populations. We demonstrate that substantial suppression can nonetheless be achieved if releases are deployed in a uniform spatial pattern using strains combining multiple lethal elements, illustrating the importance of detailed spatial models for guiding genetic mosquito control strategies. PMID:23284949

  6. The absence of detectable fetal microchimerism in nontransgenic goats (Capra aegagrus hircus) bearing transgenic offspring.

    PubMed

    Steinkraus, H B; Rothfuss, H; Jones, J A; Dissen, E; Shefferly, E; Lewis, R V

    2012-02-01

    Regulations for the disposal of genetically engineered animals are strict due to concern for their inappropriate introduction into the food chain, and of the possible public health and environmental impacts of these organisms. Nontransgenic animals that give birth to transgenic offspring are treated as if they are transgenic due to concern of fetal cells crossing the placental barrier and residing in the mother (fetal-maternal microchimerism). Determining whether or not fetal-fetal or fetal-maternal transfer of DNA or cells occurs during caprine gestation is critical to effectively protect the public without culling animals that pose no risk. Additionally, fetal-maternal transfer, should it exist in the goat, could contraindicate the rebreeding of nontransgenic dams due to the possible transfer of fetal cells from 1 pregnancy to the fetus of subsequent pregnancies. Fetal-maternal transfer in Capra hircus has not been reported in the literature, although it has been reported in another ruminant, Bos taurus. We examined blood from nontransgenic dams that carried transgenic offspring using a PCR method sensitive enough to detect the presence of a spider silk transgene to a 1:100,000 dilution. At this sensitivity, we did not detect the occurrence of fetal-maternal transfer in 5 nontransgenic dams. Likewise, fetal-fetal transfer was not observed from a transgenic to a nontransgenic twin in utero. To test tissue-specific expression of the silk transgene, proteins purified from standard necropsy tissue from a lactating transgenic dam were examined by Western blot analysis. Silk protein expression was only observed in mammary tissue consistent with the tissue specificity of the β-casein promoter used in the transgenic construct. We report evidence collected from a limited caprine breeding pool against transfer of transgenes in utero from fetus to dam and fetus to fetus. In addition, we show evidence that the β-casein promoter in our expression construct is not expressed

  7. Production of transgenic dairy goat expressing human α-lactalbumin by somatic cell nuclear transfer.

    PubMed

    Feng, Xiujing; Cao, Shaoxian; Wang, Huili; Meng, Chunhua; Li, Jingxin; Jiang, Jin; Qian, Yong; Su, Lei; He, Qiang; Zhang, Qingxiao

    2015-02-01

    Production of human α-lactalbumin (hα-LA) transgenic cloned dairy goats has great potential in improving the nutritional value and perhaps increasing the yield of dairy goat milk. Here, a mammary-specific expression vector 5A, harboring goat β-lactoglobulin (βLG) promoter, the hα-LA gene, neo(r) and EGFP dual markers, was constructed. Then, it was effectively transfected into goat mammary epithelial cells (GMECs) and the expression of hα-LA was investigated. Both the hα-LA transcript and protein were detected in the transfected GMECs after the induction of hormonal signals. In addition, the 5A vector was introduced into dairy goat fetal fibroblasts (transfection efficiency ≈60-70%) to prepare competent transgenic donor cells. A total of 121 transgenic fibroblast clones were isolated by 96-well cell culture plates and screened with nested-PCR amplification and EGFP fluorescence. After being frozen for 8 months, the transgenic cells still showed high viabilities, verifying their ability as donor cells. Dairy goat cloned embryos were produced from these hα-LA transgenic donor cells by somatic cell nuclear transfer (SCNT), and the rates of fusion, cleavage, and the development to blastocyst stages were 81.8, 84.4, and 20.0%, respectively. A total of 726 reconstructed embryos derived from the transgenic cells were transferred to 74 recipients and pregnancy was confirmed at 90 days in 12 goats. Of six female kids born, two carried hα-LA and the hα-LA protein was detected in their milk. This study provides an effective system to prepare SCNT donor cells and transgenic animals for human recombinant proteins.

  8. Development of transgenic strains for the biological control of the Mexican fruit fly, Anastrepha ludens.

    PubMed

    Meza, J Salvador; Nirmala, Xavier; Zimowska, Grazyna J; Zepeda-Cisneros, C Silvia; Handler, Alfred M

    2011-01-01

    The Mexican fruit fly, Anastrepha ludens, is a highly significant agricultural pest species that has been genetically transformed with a piggyBac-based transposon vector system using independent vector and transposase helper plasmids. Minimum estimated germ-line transformation frequencies were approximately 13-21% per fertile G(0) individual, similar to previously reported frequencies using single vector-helper plasmids. Two vector constructs were tested with potential importance to transgenic strain development for mexfly biological control. The first allows post-integration stabilization of a transposon-vector by deletion of a terminal sequence necessary for mobilization. The complete pB[L1-EGFP-L2-DsRed-R1] vector was integrated into the Chiapas wild type strain with subsequent deletion of the L2-DsRed-R1 sub-vector carrying the piggyBac 3' terminal sequence. Quality control tests for three of the stabilization vector lines (previous to stabilization) assessed viability at all life stages, fertility, adult flight ability, and adult male sexual competitiveness. All three transgenic lines were less fit compared to the wild strain by approximately 5-10% in most tests, however, there was no significant difference in sexual competitiveness which is the major prerequisite for optimal strain release. The second vector, pB[XL-EGFP, Asß2-tub-DsRed.T3], has the DsRed.T3 fluorescent protein reporter gene regulated by the A. suspensa Asß2-tubulin promoter, that resulted in testis and sperm-specific DsRed fluorescence in transgenic male mexflies. Fluorescent sperm bundles were unambiguously observed in the spermathecae of non-transgenic females mated to transgenic males. One transgenic line apparently had a male-specific Y-chromosome insertion, having potential use for sexing by fluorescent-embryo sorting. All transgenic lines expressed easily detectable and stable fluorescence in adults allowing their identification after trapping in the field.

  9. [Detection of transgenic crop with gene chip].

    PubMed

    Huang, Ying-Chun; Sun, Chun-Yun; Feng, Hong; Hu, Xiao-Dong; Yin, Hai-Bin

    2003-05-01

    Some selected available sequences of reporter genes,resistant genes, promoters and terminators are amplified by PCR for the probes of transgenic crop detection gene chip. These probes are arrayed at definite density and printed on the surface of amino-slides by bioRobot MicroGrid II. Results showed that gene chip worked quickly and correctly, when transgenic rice, pawpaw,maize and soybean were applied.

  10. Transgenic animals resistant to infectious diseases.

    PubMed

    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.

  11. Transgenic Wheat, Barley and Oats: Future Prospects

    NASA Astrophysics Data System (ADS)

    Dunwell, Jim M.

    Following the success of transgenic maize and rice, methods have now been developed for the efficient introduction of genes into wheat, barley and oats. This review summarizes the present position in relation to these three species, and also uses information from field trial databases and the patent literature to assess the future trends in the exploitation of transgenic material. This analysis includes agronomic traits and also discusses opportunities in expanding areas such as biofuels and biopharming.

  12. Carrie Chapman Catt and Woman Suffrage.

    ERIC Educational Resources Information Center

    Hardesty, Carolyn, Ed.

    1989-01-01

    Most of the material for this issue of the "Goldfinch," which explores the life of Carrie Chapman Catt, came from the archives of the State Historical Society of Iowa. Carrie Chapman Catt (1859-1947) was an Iowan who advocated woman suffrage and spent 26 years actively working for that cause. The issue contains a biography of Catt, and…

  13. Transgene flow: Facts, speculations and possible countermeasures

    PubMed Central

    Ryffel, Gerhart U

    2014-01-01

    Convincing evidence has accumulated that unintended transgene escape occurs in oilseed rape, maize, cotton and creeping bentgrass. The escaped transgenes are found in variant cultivars, in wild type plants as well as in hybrids of sexually compatible species. The fact that in some cases stacked events are present that have not been planted commercially, implies unintended recombination of transgenic traits. As the consequences of this continuous transgene escape for the ecosystem cannot be reliably predicted, I propose to use more sophisticated approaches of gene technology in future. If possible GM plants should be constructed using either site-directed mutagenesis or cisgenic strategies to avoid the problem of transgene escape. In cases where a transgenic trait is needed, efficient containment should be the standard approach. Various strategies available or in development are discussed. Such a cautious approach in developing novel types of GM crops will enhance the sustainable potential of GM crops and thus increase the public trust in green gene technology. PMID:25523171

  14. The TGV transgenic vectors for single-copy gene expression from the Escherichia coli chromosome.

    PubMed

    Gumbiner-Russo, L M; Lombardo, M J; Ponder, R G; Rosenberg, S M

    2001-07-25

    Plasmid-based cloning and expression of genes in Escherichia coli can have several problems: plasmid destabilization; toxicity of gene products; inability to achieve complete repression of gene expression; non-physiological overexpression of the cloned gene; titration of regulatory proteins; and the requirement for antibiotic selection. We describe a simple system for cloning and expression of genes in single copy in the E. coli chromosome, using a non-antibiotic selection for transgene insertion. The transgene is inserted into a vector containing homology to the chromosomal region flanking the attachment site for phage lambda. This vector is then linearized and introduced into a recombination-proficient E. coli strain carrying a temperature-sensitive lambda prophage. Selection for replacement of the prophage with the transgene is performed at high temperature. Once in the chromosome, transgenes can be moved into other lysogenic E. coli strains using standard phage-mediated transduction techniques, selecting against a resident prophage. Additional vector constructs provide an arabinose-inducible promoter (P(BAD)), P(BAD) plus a translation-initiation sequence, and optional chloramphenicol-, tetracycline-, or kanamycin-resistance cassettes. These Transgenic E. coli Vectors (TGV) allow drug-free, single-copy expression of genes from the E. coli chromosome, and are useful for genetic studies of gene function.

  15. Stacking of antimicrobial genes in potato transgenic plants confers increased resistance to bacterial and fungal pathogens.

    PubMed

    Rivero, Mercedes; Furman, Nicolás; Mencacci, Nicolás; Picca, Pablo; Toum, Laila; Lentz, Ezequiel; Bravo-Almonacid, Fernando; Mentaberry, Alejandro

    2012-01-20

    Solanum tuberosum plants were transformed with three genetic constructions expressing the Nicotiana tabacum AP24 osmotine, Phyllomedusa sauvagii dermaseptin and Gallus gallus lysozyme, and with a double-transgene construction expressing the AP24 and lysozyme sequences. Re-transformation of dermaseptin-transformed plants with the AP24/lysozyme construction allowed selection of plants simultaneously expressing the three transgenes. Potato lines expressing individual transgenes or double- and triple-transgene combinations were assayed for resistance to Erwinia carotovora using whole-plant and tuber infection assays. Resistance levels for both infection tests compared consistently for most potato lines and allowed selection of highly resistant phenotypes. Higher resistance levels were found in lines carrying the dermaseptin and lysozyme sequences, indicating that theses proteins are the major contributors to antibacterial activity. Similar results were obtained in tuber infection tests conducted with Streptomyces scabies. Plant lines showing the higher resistance to bacterial infections were challenged with Phytophthora infestans, Rhizoctonia solani and Fusarium solani. Considerable levels of resistance to each of these pathogens were evidenced employing semi-quantitative tests based in detached-leaf inoculation, fungal growth inhibition and in vitro plant inoculation. On the basis of these results, we propose that stacking of these transgenes is a promising approach to achieve resistance to both bacterial and fungal pathogens.

  16. Cre/ lox site-specific recombination controls the excision of a transgene from the rice genome.

    PubMed

    Hoa, T. T. C.; Bong, B. B.; Huq, E.; Hodges, T. K.

    2002-03-01

    The Cre/ lox site-specific recombination controls the excision of a target DNA segment by recombination between two loxsites flanking it, mediated by the Cre recombinase. We have studied the functional expression of the Cre/ lox system to excise a transgene from the rice genome. We developed transgenic plants carrying the target gene, hygromycin phosphotransferase ( hpt) flanked by two lox sites and transgenic plants harboring the Cre gene. Each lox plant was crossed with each Cre plant reciprocally. In the Cre /lox hybrid plants, the Cre recombinase mediates recombination between two lox sites, resulting in excision of the hpt gene. The recombination event could be detected because it places the CaMV 35S promoter of the hpt gene adjacent to a promoterless gusA gene; as a result the gusA gene is activated and its expression could be visualized. In 73 Cre /lox hybrid plants from various crosses of T0 transgenic plants, 19 expressed GUS, and in 132 Cre /lox hybrid plants from crosses of T2 transgenic plants, 77 showed GUS expression. Molecular data proved the excision event occurred in all the GUS(+) plants. Recombination occurred with high efficiency at the early germinal stage, or randomly during somatic development stages.

  17. Gun Carrying by High School Students in Boston, MA: Does Overestimation of Peer Gun Carrying Matter?

    ERIC Educational Resources Information Center

    Hemenway, David; Vriniotis, Mary; Johnson, Renee M.; Miller, Matthew; Azrael, Deborah

    2011-01-01

    This paper investigates: (1) whether high school students overestimate gun carrying by their peers, and (2) whether those students who overestimate peer gun carrying are more likely to carry firearms. Data come from a randomly sampled survey conducted in 2008 of over 1700 high school students in Boston, MA. Over 5% of students reported carrying a…

  18. Population growth and earth's human carrying capacity.

    PubMed

    Cohen, J E

    1995-07-21

    Earth's capacity to support people is determined both by natural constraints and by human choices concerning economics, environment, culture (including values and politics), and demography. Human carrying capacity is therefore dynamic and uncertain. Human choice is not captured by ecological notions of carrying capacity that are appropriate for nonhuman populations. Simple mathematical models of the relation between human population growth and human carrying capacity can account for faster-than-exponential population growth followed by a slowing population growth rate, as observed in recent human history.

  19. Carry on: spontaneous object carrying in 13-month-old crawling and walking infants.

    PubMed

    Karasik, Lana B; Adolph, Karen E; Tamis-LeMonda, Catherine S; Zuckerman, Alyssa L

    2012-03-01

    Carrying objects requires coordination of manual action and locomotion. This study investigated spontaneous carrying in 24 walkers who were 13 months old and 26 crawlers who were 13 months old during 1-hr, naturalistic observations in the infants' homes. Carrying was more common in walkers, but crawlers also carried objects. Typically, walkers carried objects in their hands, whereas crawlers multitasked by using their hands simultaneously for holding objects and supporting their bodies. Locomotor experience predicted frequency of carrying in both groups, suggesting that experienced crawlers and walkers perceive their increased abilities to handle objects while in motion. Despite additional biomechanical constraints imposed by holding an object, carrying may actually improve upright balance: Crawlers rarely fell while carrying an object, and walkers were more likely to fall without an object in hand than while carrying. Thus, without incurring an additional risk of falling, spontaneous carrying may provide infants with new avenues for combining locomotor and manual skills and for interacting with their environments.

  20. qPCR for quantification of transgene expression and determination of transgene copy number.

    PubMed

    Fletcher, Stephen J

    2014-01-01

    Quantitative real-time PCR (qPCR) is a mature technology that can be used to accurately quantify the number of copies of a target nucleic acid in a sample. Here, we describe a method for using this technology to determine the copy number of a transgene stably integrated into a plant's genome and to ascertain the level of transgene expression.

  1. Virus-induced gene silencing in transgenic plants: transgene silencing and reactivation associate with two patterns of transgene body methylation.

    PubMed

    Zhao, Mingmin; San León, David; Delgadillo, Ma Otilia; García, Juan Antonio; Simón-Mateo, Carmen

    2014-08-01

    We used bisulfite sequencing to study the methylation of a viral transgene whose expression was silenced upon plum pox virus infection of the transgenic plant and its subsequent recovery as a consequence of so-called virus-induced gene silencing (VIGS). VIGS was associated with a general increase in the accumulation of small RNAs corresponding to the coding region of the viral transgene. After VIGS, the transgene promoter was not methylated and the coding region showed uneven methylation, with the 5' end being mostly unmethylated in the recovered tissue or mainly methylated at CG sites in regenerated silenced plants. The methylation increased towards the 3' end, which showed dense methylation in all three contexts (CG, CHG and CHH). This methylation pattern and the corresponding silenced status were maintained after plant regeneration from recovered silenced tissue and did not spread into the promoter region, but were not inherited in the sexual offspring. Instead, a new pattern of methylation was observed in the progeny plants consisting of disappearance of the CHH methylation, similar CHG methylation at the 3' end, and an overall increase in CG methylation in the 5' end. The latter epigenetic state was inherited over several generations and did not correlate with transgene silencing and hence virus resistance. These results suggest that the widespread CG methylation pattern found in body gene bodies located in euchromatic regions of plant genomes may reflect an older silencing event, and most likely these genes are no longer silenced.

  2. Effects of carrying methods and box handles on two-person team carrying capacity for females.

    PubMed

    Wu, Swei-Pi; Chang, Shu-Yu

    2010-07-01

    This study used a psychophysical approach to examine the effects of carrying methods and the presence or absence of box handles on the maximum acceptable weight carried and resulting responses (heart rate and rating of perceived exertion) in a two-person carrying task. After training, 16 female subjects performed a two-person carrying task at knuckle height for an 8-h work period. Each subject performed 4 different carrying combinations two times. The independent variables were carrying methods (parallel and tandem walking) and box handles (with and without handles). For comparison with two-person carrying, the subjects also performed one-person carrying. The results showed that the maximum acceptable weight carried (MAWC), heart rate (HR), and rating of perceived exertion (RPE) were significantly affected by the presence of box handles. However, the subjects' MAWC, HR, and RPE values were not significantly influenced by the carrying methods. The test-retest reliability of the psychophysical approach was 0.945. The carrying efficiency of two-person carrying was 96.2% of the one-person carrying method. In general, the use of box with handles allows the subjects to carry a higher MAWC (with lower HR and RPE) compared to carrying boxes without handles.

  3. 7 CFR 1437.402 - Carrying capacity.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... Determining Coverage of Forage Intended for Animal Consumption § 1437.402 Carrying capacity. (a) CCC will... management and maintenance practices are improvements over those practices generally associated with...

  4. CCP: Sierra Nevada Captive-Carry Test

    NASA Video Gallery

    Sierra Nevada Corporation (SNC) Space System's Dream Chaser design passed one of its most complex tests to date with a successful captive-carry test conducted near the Rocky Mountain Metropolitan A...

  5. Infections That Pets Carry (For Parents)

    MedlinePlus

    ... eczema should probably avoid aquariums. continue Dogs and Cats Dogs and cats are popular pets but can carry infections such ... be in the intestinal tract of infected dogs, cats, hamsters, birds, and certain farm animals. A person ...

  6. A Primer for Using Transgenic Insecticidal Cotton in Developing Countries

    PubMed Central

    Showalter, Ann M.; Heuberger, Shannon; Tabashnik, Bruce E.; Carrière, Yves

    2009-01-01

    Many developing countries face the decision of whether to approve the testing and commercial use of insecticidal transgenic cotton and the task of developing adequate regulations for its use. In this review, we outline concepts and provide information to assist farmers, regulators and scientists in making decisions concerning this technology. We address seven critical topics: 1) molecular and breeding techniques used for the development of transgenic cotton cultivars, 2) properties of transgenic cotton cultivars and their efficacy against major insect pests, 3) agronomic performance of transgenic cotton in developing countries, 4) factors affecting transgene expression, 5) impact of gene flow between transgenic and non-transgenic cotton, 6) non-target effects of transgenic cotton, and 7) management of pest resistance to transgenic cotton. PMID:19613464

  7. Transgene and mitochondrial DNA are indicators of efficient composting of transgenic pig carcasses.

    PubMed

    Murray, Dave; Meidinger, Roy G; Golovan, Serguei P; Phillips, John P; O'Halloran, Ivan P; Fan, Ming Z; Hacker, Roger R; Forsberg, Cecil W

    2007-07-01

    Composting is an environmentally sound method for the disposal of on-farm livestock mortalities that generates material suitable for use as fertilizer; however, this method is not generally permitted for disposal of transgenic livestock mortalities during the research and development phase. This study has explored the application of the polymerase chain reaction (PCR) as a method for assessing the persistence of transgene and mitochondrial DNA markers during the composting of euthanized transgenic pig. There was at least a 10(7) fold reduction of genetic material to a level that not either transgene or mitochondrion markers were detectable. At the end of the composting period, only bone fragments that were completely demineralised and chalky were detected. Chemically the compost was similar to that from pig litter and poultry mortalities, except the copper content was lower. Based on these data, composting appears to be an appropriate method for the disposal of transgenic animals.

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

    PubMed

    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.

  9. Mechanical analysis of infant carrying in hominoids

    NASA Astrophysics Data System (ADS)

    Amaral, Lia Q.

    2008-04-01

    In all higher nonhuman primates, species survival depends upon safe carrying of infants clinging to body hair of adults. In this work, measurements of mechanical properties of ape hair (gibbon, orangutan, and gorilla) are presented, focusing on constraints for safe infant carrying. Results of hair tensile properties are shown to be species-dependent. Analysis of the mechanics of the mounting position, typical of heavier infant carrying among African apes, shows that both clinging and friction are necessary to carry heavy infants. As a consequence, a required relationship between infant weight, hair-hair friction coefficient, and body angle exists. The hair-hair friction coefficient is measured using natural ape skin samples, and dependence on load and humidity is analyzed. Numerical evaluation of the equilibrium constraint is in agreement with the knuckle-walking quadruped position of African apes. Bipedality is clearly incompatible with the usual clinging and mounting pattern of infant carrying, requiring a revision of models of hominization in relation to the divergence between apes and hominins. These results suggest that safe carrying of heavy infants justify the emergence of biped form of locomotion. Ways to test this possibility are foreseen here.

  10. Mechanical analysis of infant carrying in hominoids

    PubMed Central

    2007-01-01

    In all higher nonhuman primates, species survival depends upon safe carrying of infants clinging to body hair of adults. In this work, measurements of mechanical properties of ape hair (gibbon, orangutan, and gorilla) are presented, focusing on constraints for safe infant carrying. Results of hair tensile properties are shown to be species-dependent. Analysis of the mechanics of the mounting position, typical of heavier infant carrying among African apes, shows that both clinging and friction are necessary to carry heavy infants. As a consequence, a required relationship between infant weight, hair–hair friction coefficient, and body angle exists. The hair–hair friction coefficient is measured using natural ape skin samples, and dependence on load and humidity is analyzed. Numerical evaluation of the equilibrium constraint is in agreement with the knuckle-walking quadruped position of African apes. Bipedality is clearly incompatible with the usual clinging and mounting pattern of infant carrying, requiring a revision of models of hominization in relation to the divergence between apes and hominins. These results suggest that safe carrying of heavy infants justify the emergence of biped form of locomotion. Ways to test this possibility are foreseen here. PMID:18030438

  11. Mechanical analysis of infant carrying in hominoids.

    PubMed

    Amaral, Lia Q

    2008-04-01

    In all higher nonhuman primates, species survival depends upon safe carrying of infants clinging to body hair of adults. In this work, measurements of mechanical properties of ape hair (gibbon, orangutan, and gorilla) are presented, focusing on constraints for safe infant carrying. Results of hair tensile properties are shown to be species-dependent. Analysis of the mechanics of the mounting position, typical of heavier infant carrying among African apes, shows that both clinging and friction are necessary to carry heavy infants. As a consequence, a required relationship between infant weight, hair-hair friction coefficient, and body angle exists. The hair-hair friction coefficient is measured using natural ape skin samples, and dependence on load and humidity is analyzed. Numerical evaluation of the equilibrium constraint is in agreement with the knuckle-walking quadruped position of African apes. Bipedality is clearly incompatible with the usual clinging and mounting pattern of infant carrying, requiring a revision of models of hominization in relation to the divergence between apes and hominins. These results suggest that safe carrying of heavy infants justify the emergence of biped form of locomotion. Ways to test this possibility are foreseen here.

  12. Effects of transgenic rootstocks on growth and development of non-transgenic scion cultivars in apple.

    PubMed

    Smolka, Anders; Li, Xue-Yuan; Heikelt, Catrin; Welander, Margareta; Zhu, Li-Hua

    2010-12-01

    Although cultivation of genetic modified (GM) annual crops has been steadily increasing in the recent 10 years, the commercial cultivation of GM fruit tree is still very limited and reports of field trials on GM fruit trees are rare. This is probably because development and evaluation of GM fruit trees require a long period of time due to long life cycles of trees. In this study, we report results from a field trial on three rolB transgenic dwarfing apple rootstocks of M26 and M9 together with non-transgenic controls grafted with five non-transgenic scion cultivars. We intended to investigate the effects of transgenic rootstock on non-transgenic scion cultivars under natural conditions as well as to evaluate the potential value of using the rolB gene to modify difficult-to-root rootstocks of fruit trees. The results showed that all rolB transgenic rootstocks significantly reduced vegetative growth including tree height regardless of scion cultivar, compared with the non-transgenic rootstocks. Flowering and fruiting were also decreased for cultivars grown on the transgenic rootstocks in most cases, but the fruit quality was not clearly affected by the transgenic rootstocks. Cutting experiment and RT-PCR analysis showed that the rolB gene was stably expressed under field conditions. PCR and RT-PCR analyses displayed that the rolB gene or its mRNA were not detectable in the scion cultivars, indicating no translocation of the transgene or its mRNA from rootstock to scion. Our results suggest that rolB modified rootstocks should be used in combination with vigorous scion cultivars in order to obtain sufficient vegetative growth and good yield. Alternatively, the rolB gene could be used to dwarf vigorous rootstocks of fruit trees or produce bonzai plants as it can significantly reduce the vegetative growth of plants.

  13. Gun carrying by high school students in Boston, MA: does overestimation of peer gun carrying matter?

    PubMed

    Hemenway, David; Vriniotis, Mary; Johnson, Renee M; Miller, Matthew; Azrael, Deborah

    2011-10-01

    This paper investigates: (1) whether high school students overestimate gun carrying by their peers, and (2) whether those students who overestimate peer gun carrying are more likely to carry firearms. Data come from a randomly sampled survey conducted in 2008 of over 1,700 high school students in Boston, MA. Over 5% of students reported carrying a gun, 9% of boys and 2% of girls. Students substantially overestimated the percentage of their peers who carried guns; the likelihood that a respondent carried a gun was strongly associated with their perception of the level of peer gun carrying. Most respondents believed it was easier for other youth to obtain guns than it was for them. Social marketing campaigns designed to lower young people's perceptions about the prevalence of peer gun carrying may be a promising strategy for reducing actual gun carrying among youth.

  14. Effects of box handle position and carrying range on bi-manual carrying capacity for females.

    PubMed

    Wu, Swei-Pi; Loiu, Yi; Chien, Te Hong

    2015-01-01

    This study utilizes a psychophysical approach to examine the effects on carrying capacity for bi-manual carrying tasks involving different handle positions and carrying ranges. A total of 16 female subjects participated in the experiment in groups of two people, and each group of subjects performed the tasks in a random order with 12 different combinations of carrying task. The independent variables are handle position (upper, middle, lower) and carrying range (F-F: floor height carried to floor height, F-W: floor height carried to waist height, W-W: waist height carried to waist height, W-F: waist height carried to floor height), the dependent variable is the maximum acceptable carried weight (MAWC), heart rate (HR), and the rating of perceived exertion (RPE). The results show that the handle position has a significant effect on MAWC and overall RPE but no significant effect on HR. Carrying range has a significant effect on the MAWC and HR, but no significant effect on overall HR. The handle position and carrying range have a significant interaction on the MAWC and HR. The RPE for different body parts shows significant differences, and the hands feel the most tired. Overall, this study confirms that the lower handle position with the W-W carrying range is the best combination for a two-person carrying task.

  15. Research advances on transgenic plant vaccines.

    PubMed

    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.

  16. Possible ecological risks of transgenic organism release when transgenes affect mating success: Sexual selection and the Trojan gene hypothesis

    PubMed Central

    Muir, William M.; Howard, Richard D.

    1999-01-01

    Widespread interest in producing transgenic organisms is balanced by concern over ecological hazards, such as species extinction if such organisms were to be released into nature. An ecological risk associated with the introduction of a transgenic organism is that the transgene, though rare, can spread in a natural population. An increase in transgene frequency is often assumed to be unlikely because transgenic organisms typically have some viability disadvantage. Reduced viability is assumed to be common because transgenic individuals are best viewed as macromutants that lack any history of selection that could reduce negative fitness effects. However, these arguments ignore the potential advantageous effects of transgenes on some aspect of fitness such as mating success. Here, we examine the risk to a natural population after release of a few transgenic individuals when the transgene trait simultaneously increases transgenic male mating success and lowers the viability of transgenic offspring. We obtained relevant life history data by using the small cyprinodont fish, Japanese medaka (Oryzias latipes) as a model. Our deterministic equations predict that a transgene introduced into a natural population by a small number of transgenic fish will spread as a result of enhanced mating advantage, but the reduced viability of offspring will cause eventual local extinction of both populations. Such risks should be evaluated with each new transgenic animal before release. PMID:10570162

  17. Development of Transgenic Papaya through Agrobacterium-Mediated Transformation

    PubMed Central

    Azad, Md. Abul Kalam; Rabbani, Md. Golam; Amin, Latifah; Sidik, Nik Marzuki

    2013-01-01

    Transgenic papaya plants were regenerated from hypocotyls and immature zygotic embryo after cocultivation with Agrobacterium tumefaciens LBA-4404 carrying a binary plasmid vector system containing neomycin phosphotransferase (nptII) gene as the selectable marker and β-glucuronidase (GUS) as the reporter gene. The explants were co-cultivated with Agrobacterium tumefaciens on regeneration medium containing 500 mg/L carbenicillin + 200 mg/L cefotaxime for one week. The cocultivated explants were transferred into the final selection medium containing 500 mg/L carbenicillin + 200 mg/L cefotaxime + 50 mg/L kanamycin for callus induction as well as plant regeneration. The callus derived from the hypocotyls of Carica papaya cv. Shahi showed the highest positive GUS activities compared to Carica papaya cv. Ranchi. The transformed callus grew vigorously and formed embryos followed by transgenic plantlets successfully. The result of this study showed that the hypocotyls of C. papaya cv. Shahi and C. papaya cv. Ranchi are better explants for genetic transformation compared to immature embryos. The transformed C. papaya cv. Shahi also showed the maximum number of plant regeneration compared to that of C. papaya cv. Ranchi. PMID:24066284

  18. Transgenic wheat carrying a barley UDP-glucosyltransferase exhibit high levels of Fusarium head blight resistance by detoxifying trichothecenes

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Fusarium head blight (FHB) is a worldwide disease of wheat and barley, mainly caused by Fusarium graminearum. During infection, the fungal pathogen produces trichothecene mycotoxins, such as deoxynivalenol (DON) and nivalenol (NIV) that increase fungal virulence. Moreover, grains contaminated with t...

  19. Optimal growth trajectories with finite carrying capacity.

    PubMed

    Caravelli, F; Sindoni, L; Caccioli, F; Ududec, C

    2016-08-01

    We consider the problem of finding optimal strategies that maximize the average growth rate of multiplicative stochastic processes. For a geometric Brownian motion, the problem is solved through the so-called Kelly criterion, according to which the optimal growth rate is achieved by investing a constant given fraction of resources at any step of the dynamics. We generalize these finding to the case of dynamical equations with finite carrying capacity, which can find applications in biology, mathematical ecology, and finance. We formulate the problem in terms of a stochastic process with multiplicative noise and a nonlinear drift term that is determined by the specific functional form of carrying capacity. We solve the stochastic equation for two classes of carrying capacity functions (power laws and logarithmic), and in both cases we compute the optimal trajectories of the control parameter. We further test the validity of our analytical results using numerical simulations.

  20. Optimal growth trajectories with finite carrying capacity

    NASA Astrophysics Data System (ADS)

    Caravelli, F.; Sindoni, L.; Caccioli, F.; Ududec, C.

    2016-08-01

    We consider the problem of finding optimal strategies that maximize the average growth rate of multiplicative stochastic processes. For a geometric Brownian motion, the problem is solved through the so-called Kelly criterion, according to which the optimal growth rate is achieved by investing a constant given fraction of resources at any step of the dynamics. We generalize these finding to the case of dynamical equations with finite carrying capacity, which can find applications in biology, mathematical ecology, and finance. We formulate the problem in terms of a stochastic process with multiplicative noise and a nonlinear drift term that is determined by the specific functional form of carrying capacity. We solve the stochastic equation for two classes of carrying capacity functions (power laws and logarithmic), and in both cases we compute the optimal trajectories of the control parameter. We further test the validity of our analytical results using numerical simulations.

  1. A myelin proteolipid protein-LacZ fusion protein is developmentally regulated and targeted to the myelin membrane in transgenic mice

    PubMed Central

    1993-01-01

    Transgenic mice were generated with a fusion gene carrying a portion of the murine myelin proteolipid protein (PLP) gene, including the first intron, fused to the E. coli LacZ gene. Three transgenic lines were derived and all lines expressed the transgene in central nervous system white matter as measured by a histochemical assay for the detection of beta-galactosidase activity. PLP-LacZ transgene expression was regulated in both a spatial and temporal manner, consistent with endogenous PLP expression. Moreover, the transgene was expressed specifically in oligodendrocytes from primary mixed glial cultures prepared from transgenic mouse brains and appeared to be developmentally regulated in vitro as well. Transgene expression occurred in embryos, presumably in pre- or nonmyelinating cells, rather extensively throughout the peripheral nervous system and within very discrete regions of the central nervous system. Surprisingly, beta- galactosidase activity was localized predominantly in the myelin in these transgenic animals, suggesting that the NH2-terminal 13 amino acids of PLP, which were present in the PLP-LacZ gene product, were sufficient to target the protein to the myelin membrane. Thus, the first half of the PLP gene contains sequences sufficient to direct both spatial and temporal gene regulation and to encode amino acids important in targeting the protein to the myelin membrane. PMID:8408224

  2. Resistance of αAI-1 transgenic chickpea (Cicer arietinum) and cowpea (Vigna unguiculata) dry grains to bruchid beetles (Coleoptera: Chrysomelidae).

    PubMed

    Lüthi, Christoph; Alvarez-Alfageme, Fernando; Ehlers, Jeffrey D; Higgins, Thomas J V; Romeis, Jörg

    2013-08-01

    Dry grain legume seeds possessing αAI-1, an α-amylase inhibitor from common bean (Phaseolus vulgaris), under the control of a cotyledon-specific promoter have been shown to be highly resistant to several important bruchid pest species. One transgenic chickpea and four cowpea lines expressing αAI-1, their respective controls, as well as nine conventional chickpea cultivars were assessed for their resistance to the bruchids Acanthoscelides obtectus (Say), Callosobruchus chinensis L. and Callosobruchus maculatus F. All transgenic lines were highly resistant to both Callosobruchus species. A. obtectus, known to be tolerant to αAI-1, was able to develop in all transgenic lines. While the cotyledons of all non-transgenic cultivars were highly susceptible to all bruchids, C. chinensis and C. maculatus larvae suffered from significantly increased mortality rates inside transgenic seeds. The main factor responsible for the partial resistance in the non-transgenic cultivars was deduced to reside in the seed coat. The αAI-1 present in seeds of transgenic chickpea and cowpea lines significantly increases their resistance to two important bruchid pest species (C. chinensis and C. maculatus) essentially to immunity. To control αAI-1 tolerant bruchid species such as A. obtectus and to avoid the development of resistance to αAI-1, varieties carrying this transgene should be protected with additional control measures.

  3. Transgenic plants for phytoremediation of herbicides.

    PubMed

    Kawahigashi, Hiroyuki

    2009-04-01

    Herbicides are economically important, but the non-point pollution that they cause may disrupt the surrounding environment. Phytoremediation of herbicides has been well studied using conventional plants. Transgenic plants produced for metabolizing herbicides and long-persisting pollutants can be used for phytoremediation of foreign chemicals in contaminated soil and water. The genes involved in the metabolism of chemical compounds can be isolated from various organisms, including bacteria, fungi, plants, and animals, and these genes are then introduced into candidate plants. Transgenic plants expressing mammalian P450s and the other enzymes showed tolerance and phytoremediation activity toward target herbicides. Transgenic plants can also enhance the absorption and detoxification of pollutants, thereby aiding the phytoremediation of contaminated environments.

  4. Toxins for transgenic resistance to hemipteran pests.

    PubMed

    Chougule, Nanasaheb P; Bonning, Bryony C

    2012-06-01

    The sap sucking insects (Hemiptera), which include aphids, whiteflies, plant bugs and stink bugs, have emerged as major agricultural pests. The Hemiptera cause direct damage by feeding on crops, and in some cases indirect damage by transmission of plant viruses. Current management relies almost exclusively on application of classical chemical insecticides. While the development of transgenic crops expressing toxins derived from the bacterium Bacillus thuringiensis (Bt) has provided effective plant protection against some insect pests, Bt toxins exhibit little toxicity against sap sucking insects. Indeed, the pest status of some Hemiptera on Bt-transgenic plants has increased in the absence of pesticide application. The increased pest status of numerous hemipteran species, combined with increased prevalence of resistance to chemical insecticides, provides impetus for the development of biologically based, alternative management strategies. Here, we provide an overview of approaches toward transgenic resistance to hemipteran pests.

  5. Assured load carrying capability and capacity credit

    NASA Astrophysics Data System (ADS)

    Pape, H.

    1981-04-01

    The determination of assured load carrying capability and the capacity credit for use in planning windpowered electric generation facilities is considered. Calculation of the available capacity of thermal power plants is described and compared with calculation of available capacity for wind turbines, taking into account outages caused by the unavailability of the primary energy, wind. The assured load carrying capability of power plants is defined. An operational definition of the capacity credit of wind turbines as related to a fixed time t Epsilon T is presented and extended to the period T.

  6. Germ Cell-Specific Excision of loxP-Flanked Transgenes in Rainbow Trout Oncorhynchus mykiss.

    PubMed

    Katayama, Naoto; Kume, Sachi; Hattori-Ihara, Shoko; Sadaie, Sakiko; Hayashi, Makoto; Yoshizaki, Goro

    2016-04-01

    Cre/loxP-mediated DNA excision in germ cell lineages could contribute substantially to the study of germ cell biology in salmonids, which are emerging as a model species in this field. However, a cell type-specific Cre/loxPsystem has not been successfully developed for any salmonid species. Therefore, we examined the feasibility of Cre/loxP-mediated, germ cell-specific gene excision and transgene activation in rainbow trout. Double-transgenic (wTg) progeny were obtained by mating a transgenic male carryingcrewith a transgenic female carrying thehsc-LRLGgene;crewas driven by rainbow troutvasaregulatory regions and thehsc-LRLGgene was made up of the rainbow troutheat-shock-cognate71promoter, theDsRedgene flanked by twoloxPsites, and theEgfpgene. PCR analysis, fluorescence imaging, and histological analysis revealed that excision of theloxP-flanked sequence and activation ofEgfpoccurred only in germ cells of wTg fish. However, progeny tests revealed that the excision efficiency ofloxP-flanked sequence in germ cells was low (≤3.27%). In contrast, the other wTg fish derived from two differentcre-transgenic males frequently excised theloxP-flanked sequence in germ cells (≤89.25%). Thus, we showed for the first time successful germ cell-specific transgene manipulation via the Cre/loxPsystem in rainbow trout. We anticipate that this technology will be suitable for studies of cell function through cell targeting, cell-linage tracing, and generating cell type-specific conditional gene knockouts and separately for developing sterile rainbow trout in aquaculture.

  7. Paucity and preferential suppression of transgenes in late replication domains of the D. melanogaster genome

    PubMed Central

    2010-01-01

    Background Eukaryotic genomes are organized in extended domains with distinct features intimately linking genome structure, replication pattern and chromatin state. Recently we identified a set of long late replicating euchromatic regions that are underreplicated in salivary gland polytene chromosomes of D. melanogaster. Results Here we demonstrate that these underreplicated regions (URs) have a low density of P-element and piggyBac insertions compared to the genome average or neighboring regions. In contrast, Minos-based transposons show no paucity in URs but have a strong bias to testis-specific genes. We estimated the suppression level in 2,852 stocks carrying a single P-element by analysis of eye color determined by the mini-white marker gene and demonstrate that the proportion of suppressed transgenes in URs is more than three times higher than in the flanking regions or the genomic average. The suppressed transgenes reside in intergenic, genic or promoter regions of the annotated genes. We speculate that the low insertion frequency of P-elements and piggyBacs in URs partially results from suppression of transgenes that potentially could prevent identification of transgenes due to complete suppression of the marker gene. In a similar manner, the proportion of suppressed transgenes is higher in loci replicating late or very late in Kc cells and these loci have a lower density of P-elements and piggyBac insertions. In transgenes with two marker genes suppression of mini-white gene in eye coincides with suppression of yellow gene in bristles. Conclusions Our results suggest that the late replication domains have a high inactivation potential apparently linked to the silenced or closed chromatin state in these regions, and that such inactivation potential is largely maintained in different tissues. PMID:20492674

  8. Gene-Splitting Technology: A Novel Approach for the Containment of Transgene Flow in Nicotiana tabacum

    PubMed Central

    Kong, Ning; Jia, Shi-Rong; Tang, Qiao-Ling; Wang, Zhi-Xing

    2014-01-01

    The potential impact of transgene escape on the environment and food safety is a major concern to the scientists and public. This work aimed to assess the effect of intein-mediated gene splitting on containment of transgene flow. Two fusion genes, EPSPSn-In and Ic-EPSPSc, were constructed and integrated into N. tabacum, using Agrobacterium tumefaciens-mediated transformation. EPSPSn-In encodes the first 295 aa of the herbicide resistance gene 5-enolpyruvyl shikimate-3-phosphate synthase (EPSPS) fused with the first 123 aa of the Ssp DnaE intein (In), whereas Ic-EPSPSc encodes the 36 C-terminal aa of the Ssp DnaE intein (Ic) fused to the rest of EPSPS C terminus peptide sequences. Both EPSPSn-In and Ic-EPSPSc constructs were introduced into the same N. tabacum genome by genetic crossing. Hybrids displayed resistance to the herbicide N-(phosphonomethyl)-glycine (glyphosate). Western blot analysis of protein extracts from hybrid plants identified full-length EPSPS. Furthermore, all hybrid seeds germinated and grew normally on glyphosate selective medium. The 6-8 leaf hybrid plants showed tolerance of 2000 ppm glyphosate in field spraying. These results indicated that functional EPSPS protein was reassembled in vivo by intein-mediated trans-splicing in 100% of plants. In order to evaluate the effect of the gene splitting technique for containment of transgene flow, backcrossing experiments were carried out between hybrids, in which the foreign genes EPSPSn-In and Ic-EPSPSc were inserted into different chromosomes, and non-transgenic plants NC89. Among the 2812 backcrossing progeny, about 25% (664 plantlets) displayed glyphosate resistance. These data indicated that transgene flow could be reduced by 75%. Overall, our findings provide a new and highly effective approach for biological containment of transgene flow. PMID:24915192

  9. Blocking of Plasmodium transmission by cooperative action of Cecropin A and Defensin A in transgenic Aedes aegypti mosquitoes.

    PubMed

    Kokoza, Vladimir; Ahmed, Abdouelaziz; Woon Shin, Sang; Okafor, Nwando; Zou, Zhen; Raikhel, Alexander S

    2010-05-04

    To overcome burden of mosquito-borne diseases, multiple control strategies are needed. Population replacement with genetically modified mosquitoes carrying antipathogen effector genes is one of the possible approaches for controlling disease transmission. However, transgenic mosquitoes with antipathogen phenotypes based on overexpression of a single type effector molecule are not efficient in interrupting pathogen transmission. Here, we show that co-overexpression of two antimicrobial peptides (AMP), Cecropin A, and Defensin A, in transgenic Aedes aegypti mosquitoes results in the cooperative antibacterial and antiPlasmodium action of these AMPs. The transgenic hybrid mosquitoes that overexpressed both Cecropin A and Defensin A under the control of the vitellogenin promoter exhibited an elevated resistance to Pseudomonas aeruginosa infection, indicating that these AMPs acted cooperatively against this pathogenic bacterium. In these mosquitoes infected with P. gallinaceum, the number of oocysts was dramatically reduced in midguts, and no sporozoites were found in their salivary glands when the mosquitoes were fed twice to reactivate transgenic AMP production. Infection experiments using the transgenic hybrid mosquitoes, followed by sequential feeding on naive chicken, and then naive wild-type mosquitoes showed that the Plasmodium transmission was completely blocked. This study suggests an approach in generating transgenic mosquitoes with antiPlasmodium refractory phenotype, which is coexpression of two or more effector molecules with cooperative action on the parasite.

  10. Transgenic expression of cyclin-dependent kinase 4 results in epidermal hyperplasia, hypertrophy, and severe dermal fibrosis.

    PubMed

    Miliani de Marval, P L; Gimenez-Conti, I B; LaCava, M; Martinez, L A; Conti, C J; Rodriguez-Puebla, M L

    2001-07-01

    In a previous report we have described the effects of expression of D-type cyclins in epithelial tissues of transgenic mice. To study the involvement of the D-type cyclin partner cyclin-dependent kinase 4 (CDK4) in epithelial growth and differentiation, transgenic mice were generated carrying the CDK4 gene under the control of a keratin 5 promoter. As expected, transgenic mice showed expression of CDK4 in the epidermal basal-cell layer. Epidermal proliferation increased dramatically and basal cell hyperplasia and hypertrophy were observed. The hyperproliferative phenotype of these transgenic mice was independent of D-type cyclin expression because no overexpression of these proteins was detected. CDK4 and CDK2 kinase activities increased in transgenic animals and were associated with elevated binding of p27(Kip1) to CDK4. Expression of CDK4 in the epidermis results in an increased spinous layer compared with normal epidermis, and a mild hyperkeratosis in the cornified layer. In addition to epidermal changes, severe dermal fibrosis was observed and part of the subcutaneous adipose tissue was replaced by connective tissue. Also, abnormal expression of keratin 6 associated with the hyperproliferative phenotype was observed in transgenic epidermis. This model provides in vivo evidence for the role of CDK4 as a mediator of proliferation in epithelial cells independent of D-type cyclin expression.

  11. Edible transgenic plant vaccines for different diseases.

    PubMed

    Jain, Aakanchha; Saini, Vinay; Kohli, Dharm Veer

    2013-01-01

    Edible plant vaccines are immunogenic preparations containing antigenic proteins rather than pathogens, therefore, they sanctify situation where there is a possibility of resurgence of disease when the antigenic preparation contains the organism in any form whatsoever. Expression of antigens as vaccines and of antibodies against antigens of pathogens in transgenic plants is a convenient and inexpensive source for various bacterial, viral, helminths, protozoan and autoimmune diseases with lower capital costs. This review describes various diseases along with the production of edible transgenic plant vaccines/proteins for the same. Thus, substituting and improvising conventional immunization methods.

  12. Generation of BAC transgenic epithelial organoids.

    PubMed

    Schwank, Gerald; Andersson-Rolf, Amanda; Koo, Bon-Kyoung; Sasaki, Nobuo; Clevers, Hans

    2013-01-01

    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.

  13. Lorentz Contraction and Current-Carrying Wires

    ERIC Educational Resources Information Center

    van Kampen, Paul

    2008-01-01

    The force between two parallel current-carrying wires is investigated in the rest frames of the ions and the electrons. A straightforward Lorentz transformation shows that what appears as a purely magnetostatic force in the ion frame appears as a combined magnetostatic and electrostatic force in the electron frame. The derivation makes use of a…

  14. Must-Carry and Public Broadcasting.

    ERIC Educational Resources Information Center

    Davenport, Elizabeth K.

    Because of the United States Court of Appeal's ruling ("Quincy Cable TV vs. Federal Communications Commission") that government regulation of what cable television stations can broadcast violates their First Amendment rights, a number of consequences have arisen concerning what cable stations are required to broadcast (must-carry rules),…

  15. Increased carrying capacity with perennial forage kochia

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Carrying capacity can be increased on grass-dominated rangeland pastures by including perennial forage kochia (Kochia prostrata) as one of the plant components. The objectives of the study reported here were to compare the differences of traditional winter pastures versus pastures with forage kochi...

  16. Carry Hard ICBM basing: A technical assessment

    SciTech Connect

    Harvey, J.R.; Schaffer, A.B.; Speed, R.; Todaro, A.F.

    1989-11-15

    Carry Hard is a deceptive, multiple-aimpoint ICBM basing concept in which hardened, encapsulated missiles are shuttled among several thousand, low-cost, water-filled vertical shelters. Since most of the essential launch and operational support equipment is carried with the missile (not provided with each shelter), the overall system costs are reduced. High system hardness permits relatively close shelter spacing, which in turn allows Carry Hard to be deployed on a comparatively small piece of land (a few hundred square miles) that could be removed from public access. Controlled access to the deployment area helps in maintaining concealment of the missiles among the shelters. If concealment is successfully maintained, the system is believed to be survivable against plausible Soviet threats, regardless of whether attack-warning information is received or acted upon. Thus, Carry Hard holds high promise as a feasible, affordable, and survivable means of ICBM deployment, and a high priority should be given to developing the concept to the point that an informed decision on full-scale engineering development can be made. 33 refs., 4 figs., 5 tabs.

  17. 7 CFR 1437.402 - Carrying capacity.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... acreage upward for the crop year NAP assistance is requested by: (1) Three percent when at least 1... have a positive impact on the forage's carrying capacity in the crop year NAP assistance is requested... crop year NAP assistance is requested; and (3) Greater than 5 percent when producers provide...

  18. 7 CFR 1437.402 - Carrying capacity.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... acreage upward for the crop year NAP assistance is requested by: (1) Three percent when at least 1... have a positive impact on the forage's carrying capacity in the crop year NAP assistance is requested... crop year NAP assistance is requested; and (3) Greater than 5 percent when producers provide...

  19. 7 CFR 1437.402 - Carrying capacity.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... acreage upward for the crop year NAP assistance is requested by: (1) Three percent when at least 1... have a positive impact on the forage's carrying capacity in the crop year NAP assistance is requested... crop year NAP assistance is requested; and (3) Greater than 5 percent when producers provide...

  20. 7 CFR 1437.402 - Carrying capacity.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... acreage upward for the crop year NAP assistance is requested by: (1) Three percent when at least 1... have a positive impact on the forage's carrying capacity in the crop year NAP assistance is requested... crop year NAP assistance is requested; and (3) Greater than 5 percent when producers provide...

  1. Error propagation in energetic carrying capacity models

    USGS Publications Warehouse

    Pearse, Aaron T.; Stafford, Joshua D.

    2014-01-01

    Conservation objectives derived from carrying capacity models have been used to inform management of landscapes for wildlife populations. Energetic carrying capacity models are particularly useful in conservation planning for wildlife; these models use estimates of food abundance and energetic requirements of wildlife to target conservation actions. We provide a general method for incorporating a foraging threshold (i.e., density of food at which foraging becomes unprofitable) when estimating food availability with energetic carrying capacity models. We use a hypothetical example to describe how past methods for adjustment of foraging thresholds biased results of energetic carrying capacity models in certain instances. Adjusting foraging thresholds at the patch level of the species of interest provides results consistent with ecological foraging theory. Presentation of two case studies suggest variation in bias which, in certain instances, created large errors in conservation objectives and may have led to inefficient allocation of limited resources. Our results also illustrate how small errors or biases in application of input parameters, when extrapolated to large spatial extents, propagate errors in conservation planning and can have negative implications for target populations.

  2. Lectin cDNA and transgenic plants derived therefrom

    DOEpatents

    Raikhel, Natasha V.

    2000-10-03

    Transgenic plants containing cDNA encoding Gramineae lectin are described. The plants preferably contain cDNA coding for barley lectin and store the lectin in the leaves. The transgenic plants, particularly the leaves exhibit insecticidal and fungicidal properties.

  3. Production of homozygous transgenic rainbow trout with enhanced disease resistance

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Previous studies conducted in our laboratory showed that transgenic medaka expressing cecropin B transgenes exhibited resistant characteristic to fish bacterial pathogens, Pseudomonas fluorescens and Vibrio anguillarum. To confirm whether antimicrobial peptide gene will also exhibit antibacterial an...

  4. High incidence of lung, bone, and lymphoid tumors in transgenic mice overexpressing mutant alleles of the p53 oncogene.

    PubMed Central

    Lavigueur, A; Maltby, V; Mock, D; Rossant, J; Pawson, T; Bernstein, A

    1989-01-01

    We have investigated the role of the p53 gene in oncogenesis in vivo by generating transgenic mice carrying murine p53 genomic fragments isolated from a mouse Friend erythroleukemia cell line or BALB/c mouse liver DNA. Elevated levels of p53 mRNA were detected in several tissues of two transgenic lines tested. Increased levels of p53 protein were also detected in most of the tissues analyzed by Western blotting (immunoblotting). Because both transgenes encoded p53 proteins that were antigenically distinct from wild-type p53, it was possible to demonstrate that overexpression of the p53 protein was mostly, if not entirely, due to the expression of the transgenes. Neoplasms developed in 20% of the transgenic mice, with a high incidence of lung adenocarcinomas, osteosarcomas, and lymphomas. Tissues such as ovaries that expressed the transgene at high levels were not at higher risk of malignant transformation than tissues expressing p53 protein at much lower levels. The long latent period and low penetrance suggest that overexpression of p53 alone is not sufficient to induce malignancies and that additional events are required. These observations provide direct evidence that mutant alleles of the p53 oncogene have oncogenic potential in vivo and that different cell types show intrinsic differences in susceptibility to malignant transformation by p53. Since recent data suggest that p53 may be a recessive oncogene, it is possible that the elevated tumor incidence results from functional inactivation of endogenous p53 by overexpression of the mutant transgene. The high incidence of lung and bone tumors suggests that p53 transgenic mice may provide a useful model to investigate the molecular events that underlie these malignancies in humans. Images PMID:2476668

  5. Germ-line transmission of lentiviral PGK-EGFP integrants in transgenic cattle: new perspectives for experimental embryology.

    PubMed

    Reichenbach, Myriam; Lim, Tiongti; Reichenbach, Horst-Dieter; Guengoer, Tuna; Habermann, Felix A; Matthiesen, Marieke; Hofmann, Andreas; Weber, Frank; Zerbe, Holm; Grupp, Thomas; Sinowatz, Fred; Pfeifer, Alexander; Wolf, Eckhard

    2010-08-01

    Lentiviral vectors are a powerful tool for the genetic modification of livestock species. We previously generated transgenic founder cattle with lentiviral integrants carrying enhanced green fluorescent protein (EGFP) under the control of the phosphoglycerate kinase (PGK) promoter. In this study, we investigated the transmission of LV-PGK-EGFP integrants through the female and male germ line in cattle. A transgenic founder heifer (#562, Kiki) was subjected to superovulation treatment and inseminated with semen from a non-transgenic bull. Embryos were recovered and transferred to synchronized recipient heifers, resulting in the birth of a healthy male transgenic calf expressing EGFP as detected by in vivo imaging. Semen from a transgenic founder bull (#561, Jojo) was used for in vitro fertilization (IVF) of in vitro matured (IVM) oocytes from non-transgenic cows. The rates of cleavage and development to blastocyst in vitro corresponded to 52.0 +/- 4.1 and 24.5 +/- 4.4%, respectively. Expression of EGFP was observed at blastocyst stage (day 7 after IVF) and was seen in 93.0% (281/302) of the embryos. 24 EGFP-expressing embryos were transferred to 9 synchronized recipients. Analysis of 2 embryos, flushed from the uterus on day 15, two fetuses recovered on day 45, and a healthy male transgenic calf revealed consistent high-level expression of EGFP in all tissues investigated. Our study shows for the first time transmission of lentiviral integrants through the germ line of female and male transgenic founder cattle. The pattern of inheritance was consistent with Mendelian rules. Importantly, high fidelity expression of EGFP in embryos, fetuses, and offspring of founder #561 provides interesting tools for developmental studies in cattle, including interactions of gametes, embryos and fetuses with their maternal environment.

  6. Transformation of Acinetobacter sp. strain BD413 by transgenic sugar beet DNA.

    PubMed

    Gebhard, F; Smalla, K

    1998-04-01

    The ability of Acinetobacter sp. strain BD413(pFG4 delta nptII) to take up and integrate transgenic plant DNA based on homologous recombination was studied under optimized laboratory conditions. Restoration of nptII, resulting in kanamycin-resistant transformants, was observed with plasmid DNA, plant DNA, and homogenates carrying the gene nptII. Molecular analysis showed that some transformants not only restored the 317-bp deletion but also obtained additional DNA.

  7. Efficient and Rapid Development of Transgenic Hamster Models of TSEs Using a Radical New Technology

    DTIC Science & Technology

    2006-09-01

    We are breeding a flock of 50 genotyped sheep to select for the high scrapie susceptibility genotype VVRRQQ. All three lines of the mouse...transgenics carrying sheep, human, and elk PrP have been now re-derived. We have observed the first transmission of the disease from our standard scrapie ...Sheep Scrapie , TOSK, transposon 16. SECURITY CLASSIFICATION OF: 18. NUMBER OF PAGES 19a. NAME OF RESPONSIBLE PERSON USAMRMC a. REPORT U b

  8. IDENTIFICATION OF ESCAPED TRANSGENIC CREEPING BENTGRASS IN OREGON

    EPA Science Inventory

    When transgenic plants are cultivated near wild species that are sexually compatible with the crop, gene flow between the crop and wild plants is possible. A resultant concern is that transgene flow and transgene introgression within wild populations could have unintended ecologi...

  9. Benefits of transgenic insect resistance in Brassica hybrids under selection

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Field trials of transgenic crops have occasionally resulted in unintentional transgene flow to closely related species. Hybridization between transgenic cultivars and close relatives may create novel forms with potential negative outcomes for wild and weedy plant populations. We report here the outc...

  10. Proof-Carrying Code with Correct Compilers

    NASA Technical Reports Server (NTRS)

    Appel, Andrew W.

    2009-01-01

    In the late 1990s, proof-carrying code was able to produce machine-checkable safety proofs for machine-language programs even though (1) it was impractical to prove correctness properties of source programs and (2) it was impractical to prove correctness of compilers. But now it is practical to prove some correctness properties of source programs, and it is practical to prove correctness of optimizing compilers. We can produce more expressive proof-carrying code, that can guarantee correctness properties for machine code and not just safety. We will construct program logics for source languages, prove them sound w.r.t. the operational semantics of the input language for a proved-correct compiler, and then use these logics as a basis for proving the soundness of static analyses.

  11. Development of Oxygen-Carrying Compounds

    DTIC Science & Technology

    1942-10-16

    hydroxy-3-nitrobenzal)ethylene- dlimine Cobelt • 30 - ili - >, > Section Part Subseotlon I* 2-Hydroxy-3-nitro-5- methylbenzaldehyde ...Schiffes bases of 2-hydroxy-3-nitrobenzal- dehyde and of 2-hydroxy-5- methylbenzaldehyde with ethylene- dlamlne had been previously found to carry...at a much lowur temperature has not as yet been definitely ascerteined. to ’--■ ■* o - 32 - I. 2-Hydroxy-3-nltro-5- methylbenzaldehyde . The

  12. Vehicle for carrying an object of interest

    DOEpatents

    Zollinger, W. Thor; Ferrante, Todd A.

    1998-01-01

    A vehicle for carrying an object of interest across a supporting surface including a frame having opposite first and second ends; a first pair of wheels fixedly mounted on the first end of the frame; a second pair of wheels pivotally mounted on the second end of the frame; and a pair of motors borne by the frame, each motor disposed in driving relation relative to one of the pairs of wheels, the motors propelling the vehicle across the supporting surface.

  13. Relativistic Electron Wave Packets Carrying Angular Momentum

    NASA Astrophysics Data System (ADS)

    Bialynicki-Birula, Iwo; Bialynicka-Birula, Zofia

    2017-03-01

    There are important differences between the nonrelativistic and relativistic description of electron beams. In the relativistic case the orbital angular momentum quantum number cannot be used to specify the wave functions and the structure of vortex lines in these two descriptions is completely different. We introduce analytic solutions of the Dirac equation in the form of exponential wave packets and we argue that they properly describe relativistic electron beams carrying angular momentum.

  14. Vehicle for carrying an object of interest

    DOEpatents

    Zollinger, W.T.; Ferrante, T.A.

    1998-10-13

    A vehicle for carrying an object of interest across a supporting surface including a frame having opposite first and second ends; a first pair of wheels fixedly mounted on the first end of the frame; a second pair of wheels pivotally mounted on the second end of the frame; and a pair of motors borne by the frame, each motor disposed in driving relation relative to one of the pairs of wheels, the motors propelling the vehicle across the supporting surface. 8 figs.

  15. Transgenic mouse models of spinal and bulbar muscular atrophy (SBMA).

    PubMed

    Katsuno, M; Adachi, H; Inukai, A; Sobue, G

    2003-01-01

    Spinal and bulbar muscular atrophy (SBMA) is a late-onset motor neuron disease characterized by proximal muscle atrophy, weakness, contraction fasciculations, and bulbar involvement. Only males develop symptoms, while female carriers usually are asymptomatic. A specific treatment for SBMA has not been established. The molecular basis of SBMA is the expansion of a trinucleotide CAG repeat, which encodes the polyglutamine (polyQ) tract, in the first exon of the androgen receptor (AR) gene. The pathologic hallmark is nuclear inclusions (NIs) containing the mutant and truncated AR with expanded polyQ in the residual motor neurons in the brainstem and spinal cord as well as in some other visceral organs. Several transgenic (Tg) mouse models have been created for studying the pathogenesis of SBMA. The Tg mouse model carrying pure 239 CAGs under human AR promoter and another model carrying truncated AR with expanded CAGs show motor impairment and nuclear NIs in spinal motor neurons. Interestingly, Tg mice carrying full-length human AR with expanded polyQ demonstrate progressive motor impairment and neurogenic pathology as well as sexual difference of phenotypes. These models recapitulate the phenotypic expression observed in SBMA. The ligand-dependent nuclear localization of the mutant AR is found to be involved in the disease mechanism, and hormonal therapy is suggested to be a therapeutic approach applicable to SBMA.

  16. Transgenic plants protected from insect attack

    NASA Astrophysics Data System (ADS)

    Vaeck, Mark; Reynaerts, Arlette; Höfte, Herman; Jansens, Stefan; de Beuckeleer, Marc; Dean, Caroline; Zabeau, Marc; Montagu, Marc Van; Leemans, Jan

    1987-07-01

    The Gram-positive bacterium Bacillus thuringiensis produces proteins which are specifically toxic to a variety of insect species. Modified genes have been derived from bt2, a toxin gene cloned from one Bacillus strain. Transgenic tobacco plants expressing these genes synthesize insecticidal proteins which protect them from feeding damage by larvae of the tobacco hornworm.

  17. Transgenic Mouse Model of Chronic Beryllium Disease

    SciTech Connect

    Gordon, Terry

    2009-05-26

    Animal models provide powerful tools for dissecting dose-response relationships and pathogenic mechanisms and for testing new treatment paradigms. Mechanistic research on beryllium exposure-disease relationships is severely limited by a general inability to develop a sufficient chronic beryllium disease animal model. Discovery of the Human Leukocyte Antigen (HLA) - DPB1Glu69 genetic susceptibility component of chronic beryllium disease permitted the addition of this human beryllium antigen presentation molecule to an animal genome which may permit development of a better animal model for chronic beryllium disease. Using FVB/N inbred mice, Drs. Rubin and Zhu, successfully produced three strains of HLA-DPB1 Glu 69 transgenic mice. Each mouse strain contains a haplotype of the HLA-DPB1 Glu 69 gene that confers a different magnitude of odds ratio (OR) of risk for chronic beryllium disease: HLA-DPB1*0401 (OR = 0.2), HLA-DPB1*0201 (OR = 15), HLA-DPB1*1701 (OR = 240). In addition, Drs. Rubin and Zhu developed transgenic mice with the human CD4 gene to permit better transmission of signals between T cells and antigen presenting cells. This project has maintained the colonies of these transgenic mice and tested the functionality of the human transgenes.

  18. Viable transgenic goats derived from skin cells.

    PubMed

    Behboodi, Esmail; Memili, Erdogan; Melican, David T; Destrempes, Margaret M; Overton, Susan A; Williams, Jennifer L; Flanagan, Peter A; Butler, Robin E; Liem, Hetty; Chen, Li How; Meade, Harry M; Gavin, William G; Echelard, Yann

    2004-06-01

    The current study was undertaken to evaluate the possibility of expanding transgenic goat herds by means of somatic cell nuclear transfer (NT) using transgenic goat cells as nucleus donors. Skin cells from adult, transgenic goats were first synchronized at quiescent stage (G0) by serum starvation and then induced to exit G0 and proceed into G1. Oocytes collected from superovulated donors were enucleated, karyoplast-cytoplast couplets were constructed, and then fused and activated simultaneously by a single electrical pulse. Fused couplets were either co-cultured with oviductal cells in TCM-199 medium (in vitro culture) or transferred to intermediate recipient goat oviducts (in vivo culture) until final transfer. The resulting morulae and blastocysts were transferred to the final recipients. Pregnancies were confirmed by ultrasonography 25-30 days after embryo transfer. In vitro cultured NT embryos developed to morulae and blastocyst stages but did not produce any pregnancies while 30% (6/20) of the in vivo derived morulae and blastocysts produced pregnancies. Two of these pregnancies were resorbed early in gestation. Of the four recipients that maintained pregnancies to term, two delivered dead fetuses 2-3 days after their due dates, and two recipients gave birth to healthy kids at term. Fluorescence in situ hybridization (FISH) analysis confirmed that both kids were transgenic and had integration sites consistent with those observed in the adult cell line.

  19. Transgenic plants with increased calcium stores

    NASA Technical Reports Server (NTRS)

    Wyatt, Sarah (Inventor); Tsou, Pei-Lan (Inventor); Robertson, Dominique (Inventor); Boss, Wendy (Inventor)

    2004-01-01

    The present invention provides transgenic plants over-expressing a transgene encoding a calcium-binding protein or peptide (CaBP). Preferably, the CaBP is a calcium storage protein and over-expression thereof does not have undue adverse effects on calcium homeostasis or biochemical pathways that are regulated by calcium. In preferred embodiments, the CaBP is calreticulin (CRT) or calsequestrin. In more preferred embodiments, the CaBP is the C-domain of CRT, a fragment of the C-domain, or multimers of the foregoing. In other preferred embodiments, the CaBP is localized to the endoplasmic reticulum by operatively associating the transgene encoding the CaBP with an endoplasmic reticulum localization peptide. Alternatively, the CaBP is targeted to any other sub-cellular compartment that permits the calcium to be stored in a form that is biologically available to the plant. Also provided are methods of producing plants with desirable phenotypic traits by transformation of the plant with a transgene encoding a CaBP. Such phenotypic traits include increased calcium storage, enhanced resistance to calcium-limiting conditions, enhanced growth and viability, increased disease and stress resistance, enhanced flower and fruit production, reduced senescence, and a decreased need for fertilizer production. Further provided are plants with enhanced nutritional value as human food or animal feed.

  20. Assessing the value of transgenic crops.

    PubMed

    Lacey, Hugh

    2002-10-01

    In the current controversy about the value of transgenic crops, matters open to empirical inquiry are centrally at issue. One such matter is a key premise in a common argument (that I summarize) that transgenic crops should be considered to have universal value. The premise is that there are no alternative forms of agriculture available to enable the production of sufficient food to feed the world. The proponents of agroecology challenge it, claiming that agroecology provides an alternative, and they deny the claim that it is well founded on empirical evidence. It is, therefore, a matter of both social and scientific importance that this premise and the criticisms of it be investigated rigorously and empirically, so that the benefits and disadvantages of transgenic-intensive agriculture and agroecology can be compared in a reliable way. Conducting adequate investigation about the potential contribution of agroecology requires that the cultural conditions of its practice (and, thus, of the practices and movements of small-scale farmers in the "third world") be strengthened--and this puts the interests of investigation into tension with the socio-economic interests driving the development of transgenics. General issues about relationship between ethical argument and empirical (scientific) investigation are raised throughout the article.

  1. Monitoring transgenic plants using in vivo markers

    SciTech Connect

    Stewart, C.N. Jr.

    1996-06-01

    The gene coding for green fluorecent protein (GFP), isolated and cloned from the jellyfish Aequorea victoria, is an ideal transgene for the monitoring of any plant species. It has the ability to fluoresce without added substrate, enzyme, or cofactor; it does not introduce morphological or sexual aberrations when expressed. 7 refs., 1 fig.

  2. Metal resistance sequences and transgenic plants

    DOEpatents

    Meagher, Richard Brian; Summers, Anne O.; Rugh, Clayton L.

    1999-10-12

    The present invention provides nucleic acid sequences encoding a metal ion resistance protein, which are expressible in plant cells. The metal resistance protein provides for the enzymatic reduction of metal ions including but not limited to divalent Cu, divalent mercury, trivalent gold, divalent cadmium, lead ions and monovalent silver ions. Transgenic plants which express these coding sequences exhibit increased resistance to metal ions in the environment as compared with plants which have not been so genetically modified. Transgenic plants with improved resistance to organometals including alkylmercury compounds, among others, are provided by the further inclusion of plant-expressible organometal lyase coding sequences, as specifically exemplified by the plant-expressible merB coding sequence. Furthermore, these transgenic plants which have been genetically modified to express the metal resistance coding sequences of the present invention can participate in the bioremediation of metal contamination via the enzymatic reduction of metal ions. Transgenic plants resistant to organometals can further mediate remediation of organic metal compounds, for example, alkylmetal compounds including but not limited to methyl mercury, methyl lead compounds, methyl cadmium and methyl arsenic compounds, in the environment by causing the freeing of mercuric or other metal ions and the reduction of the ionic mercury or other metal ions to the less toxic elemental mercury or other metals.

  3. Carry-over coarticulation in joint angles.

    PubMed

    Hansen, Eva; Grimme, Britta; Reimann, Hendrik; Schöner, Gregor

    2015-09-01

    Coarticulation indicates a dependence of a movement segment on a preceding segment (carry-over coarticulation) or on the segment that follows (anticipatory coarticulation). Here we study coarticulation in multidegrees of freedom human arm movements. We asked participants to transport a cylinder from a starting position to a center target and on to a final target. In this naturalistic setting, the human arm has ten degrees of freedom and is thus comfortably redundant for the task. We studied coarticulation by comparing movements between the same spatial locations that were either preceded by different end-effector paths (carry-over coarticulation) or followed by different end-effector paths (anticipatory coarticulation). We found no evidence for coarticulation at the level of the end-effector. We found very clear evidence, however, for carry-over, not for anticipatory coarticulation at the joint level. We used the concept of the uncontrolled manifold to systematically establish coarticulation as a form of motor equivalence, in which most of the difference between different movement contexts lies within the uncontrolled manifold that leaves the end-effector invariant. The findings are consistent with movement planning occurring at the level of the end-effector, and those movement plans being transformed to the joint level by a form of inverse kinematics. The observation of massive self-motion excludes an account that is solely based on a kinematic pseudo-inverse.

  4. The substantive equivalence of transgenic (Bt and Chi) and non-transgenic cotton based on metabolite profiles.

    PubMed

    Modirroosta, Bentol Hoda; Tohidfar, Masoud; Saba, Jalal; Moradi, Foad

    2014-03-01

    Compositional studies comparing transgenic with non-transgenic counterpart plants are almost universally required by governmental regulatory bodies. In the present study, two T(2) transgenic cotton lines containing chitinase (Line 11/57) and Bt lines (Line 61) were compared with non-transgenic counterpart. To do this, biochemical characteristics of leaves and seeds, including amino acids, fatty acids, carbohydrates, anions, and cations contents of the studied lines were analyzed using GC/MS, high-performance liquid chromatography (HPLC), and ion chromatography (IC) analyzers, respectively. polymerase chain reaction (PCR) and Western blot analyses confirmed the presence and expression of Chi and Bt genes in the studied transgenic lines. Although, compositional analysis of leaves contents confirmed no significant differences between transgenic and non-transgenic counterpart lines, but it was shown that glucose content of chitinase lines, fructose content of transgenic lines (Bt and chitinase) and asparagine and glutamine of chitinase lines were significantly higher than the non-transgenic counterpart plants. Both the transgenic lines (Bt and chitinase) showed significant decrease in the amounts of sodium in comparison to the non-transgenic counterpart plants. The experiments on the seeds showed that histidine, isoleucine, leucine, and phenylalanine contents of all transgenic and non-transgenic lines were the same, whereas other amino acids were significantly increased in the transgenic lines. Surprisingly, it was observed that the concentrations of stearic acid, myristic acid, oleic acid, and linoleic acid in the chitinase line were significantly different than those of non-transgenic counterpart plants, but these components were the same in both Bt line and its non-transgenic counterpart. It seems that more changes observed in the seed contents than leaves is via this point that seeds are known as metabolites storage organs, so they show greater changes in the

  5. Growth and endocrine effects of recombinant bovine growth hormone treatment in non-transgenic and growth hormone transgenic coho salmon.

    PubMed

    Raven, P A; Sakhrani, D; Beckman, B; Neregård, L; Sundström, L F; Björnsson, B Th; Devlin, R H

    2012-05-15

    To examine the relative growth, endocrine, and gene expression effects of growth hormone (GH) transgenesis vs. GH protein treatment, wild-type non-transgenic and GH transgenic coho salmon were treated with a sustained-release formulation of recombinant bovine GH (bGH; Posilac). Fish size, specific growth rate (SGR), and condition factor (CF) were monitored for 14 weeks, after which endocrine parameters were measured. Transgenic fish had much higher growth, SGR and CF than non-transgenic fish, and bGH injection significantly increased weight and SGR in non-transgenic but not transgenic fish. Plasma salmon GH concentrations decreased with bGH treatment in non-transgenic but not in transgenic fish where levels were similar to controls. Higher GH mRNA levels were detected in transgenic muscle and liver but no differences were observed in GH receptor (GHR) mRNA levels. In non-transgenic pituitary, GH and GHR mRNA levels per mg pituitary decreased with bGH dose to levels seen in transgenic salmon. Plasma IGF-I was elevated with bGH dose only in non-transgenic fish, while transgenic fish maintained an elevated level of IGF-I with or without bGH treatment. A similar trend was seen for liver IGF-I mRNA levels. Thus, bGH treatment increased fish growth and influenced feedback on endocrine parameters in non-transgenic but not in transgenic fish. A lack of further growth stimulation of GH transgenic fish suggests that these fish are experiencing maximal growth stimulation via GH pathways.

  6. Transgenic control of perforin gene expression

    SciTech Connect

    Lichtenheld, M.G.; Podack, E.R.; Levy, R.B.

    1995-03-01

    Perforin is a pore-forming effector molecule of CTL and NK cells. To characterize perforin gene expression and its transcriptional control mechanisms in vivo, expression of a cell surface tag, i.e., human CD4, was driven by 5.1 kb of the murin perforin 5{prime} flanking and promoter region in transgenic mice. Six out of seven transgenic lines expressed the perforin-tag hybrid gene at low to intermediate levels, depending on the integration site. Transgene expression occurred in all cells that physiologically are able to express perforin. At the whole organ level, significant amounts of transgenic mRNA and endogenous perforin mRNA were co-expressed in the lymphoid organs, as well as in the lung, the ileum, the oviduct/uterus, and the bone marrow. At the single cell level, the perforin tag was present on NK cells and on CD8{sup +}, as well as on CD4{sup +} cells. Also targeted were Thy-1.2{sup +} {gamma}{delta} T cells, but not Thy-1.2{sup -} {gamma}{delta} T cells, B cells, nor monocytes. During thymic T cell development, transgene expression occurred in double negative (CD4{sup -}CD8{sup -}) thymocytes and was detected at all subsequent stages, but exceeded the expression levels of the endogenous gene in the thymus. In conclusion, the analyzed perforin 5{prime} flanking and promoter region contains important cis-acting sequences that restrict perforin expression to T cells and NK cells, and therefore provides a unique tool for manipulating T cell and/or Nk cell-mediated immune responses in transgenic mice. On the other hand, the normal control of perforin gene expression involves at least one additional negative control mechanism that was not mediated by the transgenic promoter and upstream region. This control restricts perforin gene expression in thymically developing T cells and in most resting peripheral T cells, but can be released upon T cell activation. 43 refs., 7 figs., 1 tab.

  7. Transgenic Brain-Derived Neurotrophic Factor Modulates a Developing Cerebellar Inhibitory Synapse

    PubMed Central

    Bao, Shaowen; Chen, Lu; Qiao, Xiaoxi; Thompson, Richard F.

    1999-01-01

    Brain-derived neurotrophic factor (BDNF) has been shown to promote synapse formation and maturation in neurons of many brain regions, including inhibitory synapses. In the cerebellum, the Golgi cell-granule cell GABAergic synaptic responses undergo developmental transition from slow-decaying to fast-decaying kinetics, which parallels a developmental increase of GABAA receptor α6 subunit expression in the cerebellar granule cells. In culture, BDNF accelerates the expression of GABAA receptor α6 subunit expression in granule cells. Here we examined synaptic GABAA response kinetics in BDNF transgenic mice. The mutant mouse, which carries a BDNF transgene driven by a β-actin promoter, overexpresses BDNF (two- to fivefold increase compared with wild types) in all brain regions. Recordings of the spontaneous GABAA responses indicate that the decay time constant of the GABAergic responses decreases during early postnatal development; this transition is accelerated in the BDNF transgenic mouse. The amplitude of the spontaneous GABAA responses was also larger in the transgenic mouse than in the wild-type mouse. However, the frequency of the spontaneous GABAA responses were not different between the two groups. Our results suggest that BDNF may modulate GABAergic synapse maturation in the cerebellum. PMID:10492009

  8. Improved Transgenic Mouse Model for Studying HLA Class I Antigen Presentation

    PubMed Central

    Huang, Man; Zhang, Wei; Guo, Jie; Wei, Xundong; Phiwpan, Krung; Zhang, Jianhua; Zhou, Xuyu

    2016-01-01

    HLA class I (HLA-I) transgenic mice have proven to be useful models for studying human MHC-related immune responses over the last two decades. However, differences in the processing and presentation machinery between humans and mice may have profound effects on HLA-I restricted antigen presentation. In this study, we generated a novel human TAP-LMP (hTAP-LMP) gene cluster transgenic mouse model carrying an intact human TAP complex and two human immunoproteasome LMP subunits, PSMB8/PSMB9. By crossing the hTAP-LMP strain with different HLA-I transgenic mice, we found that the expression levels of human HLA-I molecules, especially the A3 supertype members (e.g., A11 and A33), were remarkably enhanced in corresponding HLA-I/hTAP-LMP transgenic mice. Moreover, we found that humanized processing and presentation machinery increased antigen presentation of HLA-A11-restricted epitopes and promoted the rapid reduction of hepatitis B virus (HBV) infection in HLA-A11/hTAP-LMP mice. Together, our study highlights that HLA-I/hTAP-LMP mice are an improved model for studying antigen presentation of HLA-I molecules and their related CTL responses. PMID:27634283

  9. Measuring gene flow from two birdsfoot trefoil (Lotus corniculatus) field trials using transgenes as tracer markers.

    PubMed

    De Marchis, F; Bellucci, M; Arcioni, S

    2003-06-01

    Genetic engineering is becoming a useful tool in the improvement of plants but concern has been expressed about the potential environmental risks of releasing genetically modified (GM) organisms into the environment. Attention has focused on pollen dispersal as a major issue in the risk assessment of transgenic crop plants. In this study, pollen-mediated dispersal of transgenes via cross-fertilization was examined. Plants of Lotus corniculatus L. transformed with either the Escherichia coli asparagine synthetase gene asnA or the beta-glucuronidase gene uidA, were used as the pollen donor. Nontransgenic plants belonging to the species L. corniculatus L., L. tenuis Waldst. and Kit. ex Willd, and L. pedunculatus Cav., were utilized as recipients. Two experimental fields were established in two areas of central Italy. Plants carrying the uidA gene were partially sterile, therefore only the asnA gene was used as a tracer marker. No transgene flow between L. corniculatus transformants and the nontransgenic L. tenuis and L. pedunculatus plants was detected. As regards nontransgenic L. corniculatus plants, in one location flow of asnA transgene was detected up to 18 m from the 1.8 m2 donor plot. In the other location, pollen dispersal occurred up to 120 m from the 14 m2 pollinating plot.

  10. Technical advance: stringent control of transgene expression in Arabidopsis thaliana using the Top10 promoter system

    NASA Technical Reports Server (NTRS)

    Love, J.; Scott, A. C.; Thompson, W. F.; Brown, C. S. (Principal Investigator)

    2000-01-01

    We show that the tightly regulated tetracycline-sensitive Top10 promoter system (Weinmann et al. Plant J. 1994, 5, 559-569) is functional in Arabidopsis thaliana. A pure breeding A. thaliana line (JL-tTA/8) was generated which expressed a chimeric fusion of the tetracycline repressor and the activation domain of Herpes simplex virus (tTA), from a single transgenic locus. Plants from this line were crossed with transgenics carrying the ER-targeted green fluorescent protein coding sequence (mGFP5) under control of the Top10 promoter sequence. Progeny from this cross displayed ER-targeted GFP fluorescence throughout the plant, indicating that the tTA-Top10 promoter interaction was functional in A. thaliana. GFP expression was repressed by 100 ng ml-1 tetracycline, an order of magnitude lower than the concentration used previously to repress expression in Nicotiana tabacum. Moreover, the level of GFP expression was controlled by varying the concentration of tetracycline in the medium, allowing a titred regulation of transgenic activity that was previously unavailable in A. thaliana. The kinetics of GFP activity were determined following de-repression of the Top10:mGFP5 transgene, with a visible ER-targeted GFP signal appearing from 24 to 48 h after de-repression.

  11. Horticultural characteristics of transgenic tobacco expressing the rolC gene from Agrobacterium rhizogenes

    SciTech Connect

    Scorza, R.; Zimmerman, T.W.; Cordts, J.M.; Footen, K.J. ); Ravelonandro, M. . Station de Pathologie Vegetale)

    1994-09-01

    Wisconsin 38 tobacco (Nicotiana tabacum L.) leaf discs were transformed with the disarmed Agrobacterium tumefaciens strain EHA 101 carrying the rolC gene from A. rhizogenes and NPT II and GUS genes. Shoots that regenerated on kanamycin-containing medium were confirmed as transgenic through GUS assays, polymerase chain reaction (PCR), Southern blot analyses, and transmission of the foreign genes through the sexual cycle. Transgenic plants were as short as half the height of control plants; were earlier flowering by up to 35 days; and had smaller leaves, shorter internodes, smaller seed capsules, fewer seeds, smaller flowers, and reduced pollen viability. The number of seed capsules, leaf number, and specific root length were similar between transgenic and control plants. Transgenic clones varied in the expression of the rolC-induced growth alterations as did the first generation of seedlings from these clones. Such differences suggested the potential for selecting for different levels of expression. Transformation with the rolC gene presents a potentially useful method of genetically modifying horticultural crops, particularly for flowering date, height, and leaf and flower size. Chemical names used: neomycin phosphotransferase (NPTII), [beta]-glucuronidase (GUS).

  12. Use of transgenic Aedes aegypti in Brazil: risk perception and assessment.

    PubMed

    Paes de Andrade, Paulo; Aragão, Francisco José Lima; Colli, Walter; Dellagostin, Odir Antônio; Finardi-Filho, Flávio; Hirata, Mario Hiroyuki; Lira-Neto, Amaro de Castro; Almeida de Melo, Marcia; Nepomuceno, Alexandre Lima; Gorgônio da Nóbrega, Francisco; Delfino de Sousa, Gutemberg; Valicente, Fernando Hercos; Zanettini, Maria Helena Bodanese

    2016-10-01

    The OX513A strain of Aedes aegypti, which was developed by the British company Oxitec, expresses a self-limiting transgene that prevents larvae from developing to adulthood. In April 2014, the Brazilian National Technical Commission on Biosafety completed a risk assessment of OX513A and concluded that the strain did not present new biological risks to humans or the environment and could be released in Brazil. At that point, Brazil became the first country to approve the unconstrained release of a genetically modified mosquito. During the assessment, the commission produced a comprehensive list of - and systematically analysed - the perceived hazards. Such hazards included the potential survival to adulthood of immature stages carrying the transgene - should the transgene fail to be expressed or be turned off by exposure to sufficient environmental tetracycline. Other perceived hazards included the potential allergenicity and/or toxicity of the proteins expressed by the gene, the potential for gene flow or increased transmission of human pathogens and the occupation of vacant breeding sites by other vector species. The Zika epidemic both elevated the perceived importance of Ae. aegypti as a vector - among policy-makers and regulators as well as the general public - and increased concerns over the release of males of the OX513A strain. We have therefore reassessed the potential hazards. We found that release of the transgenic mosquitoes would still be both safe and of great potential value in the control of diseases spread by Ae. aegypti, such as chikungunya, dengue and Zika.

  13. Transgenic studies on homeobox genes in nervous system development: spina bifida in Isl1 transgenic mice.

    PubMed

    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.

  14. Diversity of arthropod community in transgenic poplar-cotton ecosystems.

    PubMed

    Zhang, D J; Lu, Z Y; Liu, J X; Li, C L; Yang, M S

    2015-12-02

    Poplar-cotton agro-ecosystems are the main agricultural planting modes of plain cotton fields in China. Here, we performed a systematic survey of the diversity and population of arthropod communities in four different combination of poplar-cotton eco-systems, including I) non-transgenic poplar and non-transgenic cotton fields; II) non-transgenic poplar and transgenic cotton fields [Bacillus thuringiensis (Bt) cotton]; III) Bt transgenic poplar (high insect resistant strain Pb29) and non-transgenic cotton; and IV) transgenic poplar and transgenic cotton fields, over a period of 3 years. Based on the statistical methods used to investigate community ecology, the effects of transgenic ecosystems on the whole structure of the arthropod community, on the structure of arthropods in the nutritive layer, and on the similarity of arthropod communities were evaluated. The main results were as follows: the transgenic poplar-cotton ecosystem has a stronger inhibitory effect on insect pests and has no impact on the structure of the arthropod community, and therefore, maintains the diversity of the arthropod community. The character index of the community indicated that the structure of the arthropod community of the transgenic poplar-cotton ecosystem was better than that of the poplar-cotton ecosystem, and that system IV had the best structure. As for the abundance of nutritional classes, the transgenic poplar-cotton ecosystem was also better than that of the non-transgenic poplar-cotton ecosystem. The cluster analysis and similarity of arthropod communities between the four different transgenic poplar-cotton ecosystems illustrated that the structure of the arthropod community excelled in the small sample of the transgenic poplar-cotton ecosystems.

  15. Information carrying capacity of a cosmological constant

    NASA Astrophysics Data System (ADS)

    Simidzija, Petar; Martín-Martínez, Eduardo

    2017-01-01

    We analyze the exchange of information in different cosmological backgrounds when sender and receiver are timelike separated and communicate through massless fields (without the exchange of light signals). Remarkably, we show that the dominance of a cosmological constant makes the amount of recoverable information imprinted in the field by the sender extremely resilient: it does not decay in time or with the spatial separation of the sender and receiver, and it actually increases with the rate of expansion of the Universe. This is in stark contrast with the information carried by conventional light signals and with previous results on timelike communication through massless fields in matter-dominated cosmologies.

  16. Carrying Synchronous Voice Data On Asynchronous Networks

    NASA Technical Reports Server (NTRS)

    Bergman, Larry A.

    1990-01-01

    Buffers restore synchronism for internal use and permit asynchronism in external transmission. Proposed asynchronous local-area digital communication network (LAN) carries synchronous voice, data, or video signals, or non-real-time asynchronous data signals. Network uses double buffering scheme that reestablishes phase and frequency references at each node in network. Concept demonstrated in token-ring network operating at 80 Mb/s, pending development of equipment operating at planned data rate of 200 Mb/s. Technique generic and used with any LAN as long as protocol offers deterministic (or bonded) access delays and sufficient capacity.

  17. Efficient Generation of Mice with Consistent Transgene Expression by FEEST

    PubMed Central

    Gao, Lei; Jiang, Yonghua; Mu, Libing; Liu, Yanbin; Wang, Fengchao; Wang, Peng; Zhang, Aiqun; Tang, Nan; Chen, Ting; Luo, Minmin; Yu, Lei; Gao, Shaorong; Chen, Liang

    2015-01-01

    Transgenic mouse models are widely used in biomedical research; however, current techniques for producing transgenic mice are limited due to the unpredictable nature of transgene expression. Here, we report a novel, highly efficient technique for the generation of transgenic mice with single-copy integration of the transgene and guaranteed expression of the gene-of-interest (GOI). We refer to this technique as functionally enriched ES cell transgenics, or FEEST. ES cells harboring an inducible Cre gene enabled the efficient selection of transgenic ES cell clones using hygromycin before Cre-mediated recombination. Expression of the GOI was confirmed by assaying for the GFP after Cre recombination. As a proof-of-principle, we produced a transgenic mouse line containing Cre-activatable tTA (cl-tTA6). This tTA mouse model was able to induce tumor formation when crossed with a transgenic mouse line containing a doxycycline-inducible oncogene. We also showed that the cl-tTA6 mouse is a valuable tool for faithfully recapitulating the clinical course of tumor development. We showed that FEEST can be easily adapted for other genes by preparing a transgenic mouse model of conditionally activatable EGFR L858R. Thus, FEEST is a technique with the potential to generate transgenic mouse models at a genome-wide scale. PMID:26573149

  18. Glucose metabolic gene expression in growth hormone transgenic coho salmon.

    PubMed

    Panserat, Stéphane; Kamalam, Biju Sam; Fournier, Jeanne; Plagnes-Juan, Elisabeth; Woodward, Krista; Devlin, Robert H

    2014-04-01

    Salmonids are generally known to be glucose intolerant. However, previous studies have shown that growth hormone (GH) transgenic coho salmon display modified nutritional regulation of glycolysis and lipogenesis compared to non-transgenic fish, suggesting the potential for better use of glucose in GH transgenic fish. To examine this in detail, GH transgenic and non-transgenic coho salmon were subjected to glucose tolerance test and subsequent metabolic assessments. After intra-peritoneal injection of 250mg/kg glucose, we analysed post-injection kinetics of glycaemia and expression of several key target genes highly involved in glucose homeostasis in muscle and liver tissues. Our data show no significant differences in plasma glucose levels during peak hyperglycaemia (3-6h after injection), demonstrating a similar glucose tolerance between transgenic and non transgenic. However, and unrelated to the hyperglycaemic episode, GH transgenic fish return to a slightly lower basal glycaemia values 24h after injection. Correspondingly, GH transgenic fish exhibited higher mRNA levels of glucokinase (GK) and glucose-6-phosphate dehydrogenase (G6PDH) in liver, and glucose transporter (GLUT4) in muscle. These data suggest that these metabolic actors may be involved in different glucose use in GH transgenic fish, which would be expected to influence the glucose challenge response. Overall, our data demonstrate that GH transgenic coho salmon may be a pertinent animal model for further study of glucose metabolism in carnivorous fish.

  19. Metabolic rate of carrying added mass: a function of walking speed, carried mass and mass location.

    PubMed

    Schertzer, Eliran; Riemer, Raziel

    2014-11-01

    The effort of carrying additional mass at different body locations is important in ergonomics and in designing wearable robotics. We investigate the metabolic rate of carrying a load as a function of its mass, its location on the body and the subject's walking speed. Novel metabolic rate prediction equations for walking while carrying loads at the ankle, knees and back were developed based on experiments where subjects walked on a treadmill at 4, 5 or 6km/h bearing different amounts of added mass (up to 2kg per leg and 22kg for back). Compared to previously reported equations, ours are 7-69% more accurate. Results also show that relative cost for carrying a mass at a distal versus a proximal location changes with speed and mass. Contrary to mass carried on the back, mass attached to the leg cannot be modeled as an increase in body mass.

  20. Trait stacking in transgenic crops: challenges and opportunities.

    PubMed

    Que, Qiudeng; Chilton, Mary-Dell M; de Fontes, Cheryl M; He, Chengkun; Nuccio, Michael; Zhu, Tong; Wu, Yuexuan; Chen, Jeng S; Shi, Liang

    2010-01-01

    In recent years, there has been a rapid increase in the planting of transgenic crops with stacked traits. Most of these products have been formed by conventional breeding, i.e. the crossing of transgenic plant (event) containing individual transgenes with other event(s) containing single or double transgenic traits. Many biotech companies are developing stacked trait products with increasing numbers of insect and herbicide tolerance genes for controlling a broad range of insect pests and weeds. There has also been an increase in development of technologies for molecular stacking of multiple traits in a single transgene locus. In this review we look at the status of stacked trait products, crop trait stacking technologies and the technical challenges we are facing. We also review recent progress in developing technology for assembling large transgene arrays in vitro (molecular stacks), their delivery to crop plants and issues they pose for transgene expression.

  1. Co-transforming bar and CsLEA enhanced tolerance to drought and salt stress in transgenic alfalfa (Medicago sativa L.).

    PubMed

    Zhang, Jiyu; Duan, Zhen; Zhang, Daiyu; Zhang, Jianquan; Di, Hongyan; Wu, Fan; Wang, Yanrong

    2016-03-25

    Drought and high salinity are two major abiotic factors that restrict alfalfa productivity. A dehydrin protein, CsLEA, from the desert grass Cleistogenes songorica was transformed into alfalfa (Medicago sativa L.) via Agrobacterium-mediated transformation using the bar gene as a selectable marker, and the drought and salt stress tolerances of the transgenic plants were assessed. Thirty-nine of 119 transformants were positive, as screened by Basta, and further molecularly authenticated using PCR and RT-PCR. Phenotype observations revealed that the transgenic plants grew better than the wild-type (WT) plants after 15d of drought stress and 10d of salt stress: the leaves of WT alfalfa turned yellow, whereas the transgenic alfalfa leaves only wilted; after rewatering, the transgenic plants returned to a normal state, though the WT plants could not be restored. Evaluation of physiologic and biochemical indices during drought and salt stresses showed a relatively lower Na(+) content in the leaves of the transgenic plants, which would reduce toxic ion effects. In addition, the transgenic plants were able to maintain a higher relative water content (RWC), higher shoot biomass, fewer photosystem changes, decreased membrane injury, and a lower level of osmotic stress injury. These results demonstrate that overexpression of the CsLEA gene can enhance the drought and salt tolerance of transgenic alfalfa; in addition, carrying the bar gene in the genome may increase herbicide resistance.

  2. Assessment of peanut quality and compositional characteristics among transgenic sclerotinia blight-resistant and non-transgenic susceptible cultivars.

    PubMed

    Hu, Jiahuai; Telenko, Darcy E P; Phipps, Patrick M; Grabau, Elizabeth A

    2014-08-06

    This study presents the results of a comparison that includes an analysis of variance and a canonical discriminant analysis to determine compositional equivalence and similarity between transgenic, sclerotinia blight-resistant and non-transgenic, susceptible cultivars of peanut in 3 years of field trials. Three Virginia-type cultivars (NC 7, Wilson, and Perry) and their corresponding transgenic lines (N70, W73, and P39) with a barley oxalate oxidase gene were analyzed for differences in key mineral nutrients, fatty acid components, hay constituents, and grade characteristics. Results from both analyses demonstrated that transgenic lines were compositionally similar to their non-transgenic parent cultivar in all factors as well as market-grade characteristics and nutritional value. Transgenic lines expressing oxalate oxidase for resistance to sclerotinia blight were substantially equivalent to their non-transgenic parent cultivar in quality and compositional characteristics.

  3. Economic growth, carrying capacity, and the environment

    SciTech Connect

    Arrow, K.; Bolin, B.; Costanza, R.; Dasgupta, P.; Folke, C.; Maeler, K.G.; Holling, C.S.; Jansson, B.O.; Levin, S.; Perrings, C.

    1995-04-28

    National and international economic policy has usually ignored the environment. In areas where the environment is beginning to impinge on policy, as in the General Agreement on Tariffs and Trade (GATT) and the North American Free Trade Agreement (NAFTA), it remains a tangential concern, and the presumption is often made that economic growth and economic liberalization (including the liberalization of international trade) are, in some sense, good for the environment. This notion has meant that economy-wide policy reforms designed to promote growth and liberalization have been encouraged with little regard to their environmental consequences, presumably on the assumption that these consequences would either take care of themselves or could be dealt with separately. In this article, we discuss the relation between economic growth and environmental quality, and the link between economic activity and the carrying capacity and resilience of the environment.

  4. Ectopic expression of the agouti gene in transgenic mice causes obesity, features of type II diabetes, and yellow fur.

    PubMed

    Klebig, M L; Wilkinson, J E; Geisler, J G; Woychik, R P

    1995-05-23

    Mice that carry the lethal yellow (Ay) or viable yellow (Avy) mutation, two dominant mutations of the agouti (a) gene in mouse chromosome 2, exhibit a phenotype that includes yellow fur, marked obesity, a form of type II diabetes associated with insulin resistance, and an increased susceptibility to tumor development. Molecular analyses of these and several other dominant "obese yellow" a-locus mutations suggested that ectopic expression of the normal agouti protein gives rise to this complex pleiotropic phenotype. We have now tested this hypothesis directly by generating transgenic mice that ectopically express an agouti cDNA clone encoding the normal agouti protein in all tissues examined. Transgenic mice of both sexes have yellow fur, become obese, and develop hyperinsulinemia. In addition, male transgenic mice develop hyperglycemia by 12-20 weeks of age. These results demonstrate conclusively that the ectopic agouti expression is responsible for most, if not all, of the phenotypic traits of the dominant, obese yellow mutants.

  5. Overexpression and nuclear accumulation of glyceraldehyde-3-phosphate dehydrogenase in a transgenic mouse model of Huntington's disease.

    PubMed

    Senatorov, Vladimir V; Charles, Vinod; Reddy, P H; Tagle, Dan A; Chuang, De-Maw

    2003-03-01

    Huntington's disease is due to an expansion of CAG repeats in the huntingtin gene. Huntingtin interacts with several proteins including glyceraldehyde-3-phosphate dehydrogenase (GAPDH). We performed immunohistochemical analysis of GAPDH expression in the brains of transgenic mice carrying the huntingtin gene with 89 CAG repeats. In all wild-type animals examined, GAPDH was evenly distributed among the different cell types throughout the brain. In contrast, the majority of transgenic mice showed GAPDH overexpression, with the most prominent GAPDH changes observed in the caudate putamen, globus pallidus, neocortex, and hippocampal formation. Double staining for NeuN and GFAP revealed that GAPDH overexpression occurred exclusively in neurons. Nissl staining analysis of the neocortex and caudate putamen indicated 24 and 27% of cell loss in transgenic mice, respectively. Subcellular fluorescence analysis revealed a predominant increase in GAPDH immunostaining in the nucleus. Thus, we conclude that mutation of huntingtin is associated with GAPDH overexpression and nuclear translocation in discrete populations of brain neurons.

  6. [Transgenic plants as medicine production systems].

    PubMed

    Okada, Y

    1997-10-01

    Transgenic plants are emerging as an important system for the expression of many recombinant proteins, especially those intended for therapeutic purpose. The production of foreign proteins in plants has several advantages. In terms of required equipment and cost, mass production in plants is far easier to achieve than techniques involving animal cells. Successful production of several proteins in plants, including human serum albumin, haemoglobin, monoclonal antibodies, viral antigens (vaccines), enkephalin, and trichosanthin, has been reported. Particularly, the demonstration that vaccine antigens can be produced in plants in their native, immunogenic forms opens exciting possibilities for the "bio-farming" of vaccines. If the antigens are orally active, food-based "edible vaccines" could allow economical production. In this review, I will discuss the progress that has been made by several groups in what is now an expanding area of medicine research that utilizes transgenic plants.

  7. Comparison of acetaminophen toxicity in primary hepatocytes isolated from transgenic mice with different appolipoprotein E alleles.

    PubMed

    Mezera, V; Kucera, O; Moravcova, A; Peterova, E; Rousar, T; Rychtrmoc, D; Sobotka, O; Cervinkova, Z

    2015-12-01

    The nuclear factor erythroid 2-related factor 2 (Nrf2) is a transcription factor, important for combating electrophilic and oxidative stress in the liver and other organs. This encompasses detoxification of hepatotoxic drugs, including acetaminophen (APAP). Recently, an association between apolipoprotein E (ApoE) genotype and Nrf2 expression was described. We compared the toxicity of APAP on primary culture hepatocytes isolated from transgenic mice carrying two different human ApoE alleles and wild-type controls. The cells were exposed to APAP in concentrations from 0.5 to 4 mM for up to 24 hours. APAP led to a dose-dependent hepatotoxicity from 1 mM after 16 h exposure in all mice tested. The toxicity was higher in hepatocytes isolated from both transgenic strains than in wild-type controls and most pronounced in ApoE3 mice. Concurrently, there was a decline in mitochondrial membrane potential, especially in ApoE3 hepatocytes. The formation of reactive oxygen species was increased after 24 hours with 2.5 mM APAP in hepatocytes of all strains tested, with the highest increase being in the ApoE3 genotype. The activity of caspases 3 and 7 did not differ among groups and was minimal after 24 hour incubation with 4 mM APAP. We observed higher lipid accumulation in hepatocytes isolated from both transgenic strains than in wild-type controls. The expression of Nrf2-dependent genes was higher in ApoE3 than in ApoE4 hepatocytes and some of these genes were induced by APAP treatment. In conclusion, transgenic mice with ApoE4 and ApoE3 alleles displayed higher susceptibility to acute APAP toxicity in vitro than wild-type mice. Of the two transgenic genotypes tested, ApoE3 allele carriers were more prone to injury.

  8. Germline transmission of a novel rat embryonic stem cell line derived from transgenic rats.

    PubMed

    Men, Hongsheng; Bauer, Beth A; Bryda, Elizabeth C

    2012-09-20

    Germline-competent rat embryonic stem (ES) cell lines are important resources for the creation of mutant rat models using ES-cell-based gene targeting technology. The ability to isolate germline-competent ES cell lines from any rat strain, including genetically modified strains, would allow for more sophisticated genetic manipulations without extensive breeding. Sprague Dawley (SD) males carrying an enhanced green fluorescent protein (EGFP) transgene were used as the founder animals for the derivation of ES cell lines. A number of ES cell lines were established and subjected to rigorous quality control testing that included assessment of pluripotency factor expression, karyotype analysis, and pathogen/sterility testing. Two male ES cell lines, SD-Tg.EC1/Rrrc and SD-Tg.EC8/Rrrc, were injected into blastocysts recovered from a cross of Dark Agouti (DA) males with SD females. Resulting chimeric animals were bred with wild-type SD mates to verify the germline transmissibility of the ES cell lines by identifying pups carrying the ES cell line-derived EGFP transgene. While both ES cell lines gave rise to chimeric animals, only SD-Tg.EC1 was germline competent. This confirms the feasibility of deriving germline-competent ES cell lines from transgenic rat strains and provides a novel ES cell line with a stable green fluorescent protein (GFP) reporter for future genetic manipulations to create new rat models.

  9. Expression of the human granulocyte-macrophage colony stimulating factor (hGM-CSF) gene under control of the 5'-regulatory sequence of the goat alpha-S1-casein gene with and without a MAR element in transgenic mice.

    PubMed

    Burkov, I A; Serova, I A; Battulin, N R; Smirnov, A V; Babkin, I V; Andreeva, L E; Dvoryanchikov, G A; Serov, O L

    2013-10-01

    Expression of the human granulocyte-macrophage colony-stimulating factor (hGM-CSF) gene under the control of the 5'-regulatory sequence of the goat alpha-S1-casein gene with and without a matrix attachment region (MAR) element from the Drosophila histone 1 gene was studied in four and eight transgenic mouse lines, respectively. Of the four transgenic lines carrying the transgene without MAR, three had correct tissues-specific expression of the hGM-CSF gene in the mammary gland only and no signs of cell mosaicism. The concentration of hGM-CSF in the milk of transgenic females varied from 1.9 to 14 μg/ml. One line presented hGM-CSF in the blood serum, indicating ectopic expression. The values of secretion of hGM-CSF in milk of 6 transgenic lines carrying the transgene with MAR varied from 0.05 to 0.7 μg/ml, and two of these did not express hGM-CSF. Three of the four examined animals from lines of this group showed ectopic expression of the hGM-CSF gene, as determined by RT-PCR and immunofluorescence analyses, as well as the presence of hGM-CSF in the blood serum. Mosaic expression of the hGM-CSF gene in mammary epithelial cells was specific to all examined transgenic mice carrying the transgene with MAR but was never observed in the transgenic mice without MAR. The mosaic expression was not dependent on transgene copy number. Thus, the expected "protective or enhancer effect" from the MAR element on the hGM-CSF gene expression was not observed.

  10. Optimization of Biofuel Production from Transgenic Microalgae

    DTIC Science & Technology

    2008-05-31

    gene . We observed that Chlorella was resistant to a number of antibiotics that other algae were sensitive too. This necessitated a screen for the...best antibiotic resistance genes to use for transformation selectable markers. Using the Chlamydomonas ss-rbcl and psaD promoters we have obtained...transgenic Chlorella cells at high frequency that have been confirmed by PCR analysis. We also have now completed the genome sequence of the actin gene

  11. Transgenic Animal Models of Huntington's Disease.

    PubMed

    Yang, Shang-Hsun; Chan, Anthony W S

    2011-01-01

    Huntington's disease (HD) is a devastating neurodegenerative disorder that currently has no cure. In order to develop effective treatment, an understanding of HD pathogenesis and the evaluation of therapeutic efficacy of novel medications with the aid of animal models are critical steps. Transgenic animals sharing similar genetic defects that lead to HD have provided important discoveries in HD mechanisms that cell models are not able to replicate, which include psychiatric impairment, cognitive behavioral impact, and motor functions. Although transgenic HD rodent models have been widely used in HD research, it is clear that an animal model with comparable physiology to man, similar genetic defects that lead to HD, and the ability to develop similar cognitive and behavioral impairments is critical for explaining HD pathogenesis and the development of cures. Compared to HD rodents, HD transgenic nonhuman primates have not only developed comparable neuropathology but also present HD clinical features such as rigidity, seizure, dystonia, bradykinesia, and chorea that no other animal model has been able to replicate. Distinctive degenerating neurons and the accumulation of neuropil aggregates observed in HD monkey brain strongly support the hypothesis that the unique neuropathogenic events seen in HD monkey brain recapitulate HD in man. The latest development of transgenic HD primates has opened a new era of animal modeling that better represents human genetic disorders such as HD, which will accelerate the development of diagnostic tools and identifying novel biomarkers through longitudinal studies including gene expression and metabolite profiling, and noninvasive imaging. Furthermore, novel treatments with predictable efficacy in human patients can be developed using HD monkeys because of comparable neuropathology and clinical features.

  12. Transgene mobilization and regulatory uncertainty for non-GE fruit products of transgenic rootstocks.

    PubMed

    Haroldsen, Victor M; Chi-Ham, Cecilia L; Bennett, Alan B

    2012-10-31

    Genetically engineered (GE) rootstocks may offer some advantages for biotechnology applications especially in woody perennial crops such as grape or walnut. Transgrafting combines horticultural grafting practices with modern GE methods for crop improvement. Here, a non-GE conventional scion (upper stem portion) is grafted onto a transgenic GE rootstock. Thus, the scion does not contain the genetic modification present in the rootstock genome. We examined transgene presence in walnut and tomato GE rootstocks and non-GE fruit-bearing scions. Mobilization of transgene DNA, protein, and mRNA across the graft was not detected. Though transgenic siRNA mobilization was not observed in grafted tomatoes or walnut scions, transgenic siRNA signal was detected in walnut kernels. Prospective benefits from transgrafted plants include minimized risk of GE pollen flow (Lev-Yadun and Sederoff, 2001), possible use of more than one scion per approved GE rootstock which could help curb the estimated US$136 million (CropLife International, 2011) cost to bring a GE crop to international markets, as well as potential for improved consumer and market acceptance since the consumable product is not itself GE. Thus, transgrafting provides an alternative option for agricultural industries wishing to expand their biotechnology portfolio.

  13. Using empirical data to model transgene dispersal.

    PubMed Central

    Meagher, T R; Belanger, F C; Day, P R

    2003-01-01

    One element of the current public debate about genetically modified crops is that gene flow from transgenic cultivars into surrounding weed populations will lead to more problematic weeds, particularly for traits such as herbicide resistance. Evolutionary biologists can inform this debate by providing accurate estimates of gene flow potential and subsequent ecological performance of resulting hybrids. We develop a model for gene flow incorporating exponential distance and directional effects to be applied to windpollinated species. This model is applied to previously published data on gene flow in experimental plots of Agrostis stolonifera L. (creeping bentgrass), which assessed gene flow from transgenic plants resistant to the herbicide glufosinate to surrounding non-transgenic plants. Our results show that although pollen dispersal can be limited in some sites, it may be extensive in others, depending on local conditions such as exposure to wind. Thus, hybridization under field conditions is likely to occur. Given the nature of the herbicide resistance trait, we regard this trait as unlikely to persist in the absence of herbicide, and suggest that the ecological consequences of such gene flow are likely to be minimal. PMID:12831482

  14. Using empirical data to model transgene dispersal.

    PubMed

    Meagher, T R; Belanger, F C; Day, P R

    2003-06-29

    One element of the current public debate about genetically modified crops is that gene flow from transgenic cultivars into surrounding weed populations will lead to more problematic weeds, particularly for traits such as herbicide resistance. Evolutionary biologists can inform this debate by providing accurate estimates of gene flow potential and subsequent ecological performance of resulting hybrids. We develop a model for gene flow incorporating exponential distance and directional effects to be applied to windpollinated species. This model is applied to previously published data on gene flow in experimental plots of Agrostis stolonifera L. (creeping bentgrass), which assessed gene flow from transgenic plants resistant to the herbicide glufosinate to surrounding non-transgenic plants. Our results show that although pollen dispersal can be limited in some sites, it may be extensive in others, depending on local conditions such as exposure to wind. Thus, hybridization under field conditions is likely to occur. Given the nature of the herbicide resistance trait, we regard this trait as unlikely to persist in the absence of herbicide, and suggest that the ecological consequences of such gene flow are likely to be minimal.

  15. Transgenic sorghum plants via microprojectile bombardment.

    PubMed

    Casas, A M; Kononowicz, A K; Zehr, U B; Tomes, D T; Axtell, J D; Butler, L G; Bressan, R A; Hasegawa, P M

    1993-12-01

    Transgenic sorghum plants have been obtained after microprojectile bombardment of immature zygotic embryos of a drought-resistant sorghum cultivar, P898012. DNA delivery parameters were optimized based on transient expression of R and C1 maize anthocyanin regulatory elements in scutellar cells. The protocol for obtaining transgenic plants consists of the delivery of the bar gene to immature zygotic embryos and the imposition of bialaphos selection pressure at various stages during culture, from induction of somatic embryogenesis to rooting of regenerated plantlets. One in about every 350 embryos produced embryogenic tissues that survived bialaphos treatment; six transformed callus lines were obtained from three of the eight sorghum cultivars used in this research. Transgenic (T0) plants were obtained from cultivar P898012 (two independent transformation events). The presence of the bar and uidA genes in the T0 plants was confirmed by Southern blot analysis of genomic DNA. Phosphinothricin acetyltransferase activity was detected in extracts of the T0 plants. These plants were resistant to local application of the herbicide Ignite/Basta, and the resistance was inherited in T1 plants as a single dominant locus.

  16. Safety Evaluation of Transgenic Tilapia with Accelerated Growth.

    PubMed

    Guillén; Berlanga; Valenzuela; Morales; Toledo; Estrada; Puentes; Hayes; de la Fuente J

    1999-01-01

    Recent advances in modern marine biotechnology have permitted the generation of new strains of economically important fish species through the transfer of growth hormone genes. These transgenic fish strains show improved growth performance and therefore constitute a better alternative for aquaculture programs. Recently, we have obtained a transgenic tilapia line with accelerated growth. However, before introducing this line into Cuban aquaculture, environmental and food safety assessment was required by national authorities. Experiments were performed to evaluate the behavior of transgenic tilapia in comparison to wild tilapia as a way to assess the environmental impact of introducing transgenic tilapia into Cuban aquaculture. Studies were also conducted to evaluate, according to the principle of substantial equivalence, the safety of consuming transgenic tilapia as food. Behavior studies showed that transgenic tilapia had a lower feeding motivation and dominance status than controls. Food safety assessment indicated that tilapia growth hormone has no biological activity when administered to nonhuman primates. Furthermore, no effects were detected in human healthy volunteers after the consumption of transgenic tilapia. These results showed, at least under the conditions found in Cuba, no environmental implications for the introduction of this transgenic tilapia line and the safety in the consumption of tiGH-transgenic tilapia as an alternative feeding source for humans. These results support the culture and consumption of these transgenic tilapia.

  17. Field trials and tribulations--making sense of the regulations for experimental field trials of transgenic crops in Europe.

    PubMed

    Gómez-Galera, Sonia; Twyman, Richard M; Sparrow, Penelope A C; Van Droogenbroeck, Bart; Custers, René; Capell, Teresa; Christou, Paul

    2012-06-01

    Transgenic plants that are being developed for commercial cultivation must be tested under field conditions to monitor their effects on surrounding wildlife and conventional crops. Developers also use this opportunity to evaluate the performance of transgenic crops in a typical environment, although this is a matter of commercial necessity rather than regulatory compliance. Most countries have adapted existing regulations or developed new ones to deal specifically with transgenic crops and their commodities. The European Union (EU) is renowned, or perhaps notorious, for having the broadest and most stringent regulations governing such field trials in the world. This reflects its nominal adherence to the precautionary approach, which assumes all transgenic crops carry an inherent risk. Therefore, field trials in the EU need to demonstrate that the risk associated with deploying a transgenic crop has been reduced to the level where it is regarded as acceptable within the narrowly defined limits of the regulations developed and enforced (albeit inconsistently) by national and regional governments, that is, that there is no greater risk than growing an equivalent conventional crop. The involvement of national and regional competent authorities in the decision-making process can add multiple layers of bureaucracy to an already-intricate process. In this review, we use country-based case studies to show how the EU, national and regional regulations are implemented, and we propose strategies that could increase the efficiency of regulation without burdening developers with further unnecessary bureaucracy.

  18. Efficient production of omega-3 fatty acid desaturase (sFat-1)-transgenic pigs by somatic cell nuclear transfer.

    PubMed

    Pan, DengKe; Zhang, Li; Zhou, YanRong; Feng, Chong; Long, Chuan; Liu, Xiao; Wan, Rong; Zhang, Jian; Lin, AiXing; Dong, EnQiu; Wang, ShuChen; Xu, HouGang; Chen, HongXing

    2010-04-01

    Omega-3(omega-3) fatty acid desaturase transgenic pigs may improve carcass fatty acid composition. The use of transgenic pigs is also an excellent large animal model for studying the role of omega-3 fatty acids in the prevention and treatment of coronary heart disease and cancer. Transgenic pigs carrying synthesized fatty acid desaturase-1 gene (sFat-1) from Caenorhabditis briggsae by somatic cell nuclear transfer (SCNT) were produced for the first time in China. Porcine fetal fibroblast cells were transfected with a sFat-1 expression cassette by the liposome-mediated method. Transgenic embryos were reconstructed by nuclear transfer of positive cells into enucleated in vitro matured oocytes. A total of 1889 reconstructed embryos were transferred into 10 naturally cycling gilts. Nine early pregnancies were established, 7 of which went to term. Twenty-one piglets were born. The cloning efficiency was 1.1% (born piglets/transferred embryos). The integration of the sFat-1 gene was confirmed in 15 live cloned piglets by PCR and Southern blot except for 2 piglets. Expression of the sFat-1 gene in 12 of 13 piglets was detected with RT-PCR. The data demonstrates that an efficient system for sFat-1 transgenic cloned pigs was developed, which led to the successful production of piglets expressing the sFat-1 gene.

  19. Does lignin modification affect feeding preference or growth performance of insect herbivores in transgenic silver birch (Betula pendula Roth)?

    PubMed

    Tiimonen, Heidi; Aronen, Tuija; Laakso, Tapio; Saranpää, Pekka; Chiang, Vincent; Ylioja, Tiina; Roininen, Heikki; Häggman, Hely

    2005-11-01

    Transgenic silver birch (Betula pendula Roth) lines were produced in order to modify lignin biosynthesis. These lines carry COMT (caffeate/5-hydroxyferulate O-methyltransferase) gene from Populus tremuloides driven by constitutive promoter 35S CaMV (cauliflower mosaic virus) or UbB1 (ubiquitin promoter from sunflower). The decreased syringyl/guaiacyl (S/G) ratio was found in stem and leaf lignin of 35S CaMV-PtCOMT transgenic silver birch lines when compared to non-transformed control or UbB1-PtCOMT lines. In controlled feeding experiments the leaves of transgenic birch lines as well as controls were fed to insect herbivores common in boreal environment, i.e., larvae of Aethalura punctulata, Cleora cinctaria and Trichopteryx carpinata (Lepidoptera: Geometridae) as well as the adults of birch leaf-feeding beetles Agelastica alni (Coleoptera: Chrysomelidae) and Phyllobius spp. (Coleoptera: Curculionidae). The feeding preferences of these herbivores differed in some cases among the tested birch lines, but these differences could not be directly associated to lignin modification. They could as well be explained by other characteristics of leaves, either natural or caused by transgene site effects. Growth performance of lepidopteran larvae fed on transgenic or control leaves did not differ significantly.

  20. Expression of Active Fluorophore Proteins in the Milk of Transgenic Pigs Bypassing the Secretory Pathway

    PubMed Central

    Mukherjee, Ayan; Garrels, Wiebke; Talluri, Thirumala R.; Tiedemann, Daniela; Bősze, Zsuzsanna; Ivics, Zoltán; Kues, Wilfried A.

    2016-01-01

    We describe the expression of recombinant fluorescent proteins in the milk of two lines of transgenic pigs generated by Sleeping Beauty transposon-mediated genetic engineering. The Sleeping Beauty transposon consisted of an ubiquitously active CAGGS promoter driving a fluorophore cDNA, encoding either Venus or mCherry. Importantly, the fluorophore cDNAs did not encode for a signal peptide for the secretory pathway, and in previous studies of the transgenic animals a cytoplasmic localization of the fluorophore proteins was found. Unexpectedly, milk samples from lactating sows contained high levels of bioactive Venus or mCherry fluorophores. A detailed analysis suggested that exfoliated cells of the mammary epithelium carried the recombinant proteins passively into the milk. This is the first description of reporter fluorophore expression in the milk of livestock, and the findings may contribute to the development of an alternative concept for the production of bioactive recombinant proteins in the udder. PMID:27086548

  1. Transgenic rescue of phenotypic deficits in a mouse model of alternating hemiplegia of childhood.

    PubMed

    Kirshenbaum, Greer S; Dachtler, James; Roder, John C; Clapcote, Steven J

    2016-01-01

    Missense mutations in ATP1A3 encoding Na(+),K(+)-ATPase α3 are the primary cause of alternating hemiplegia of childhood (AHC). Most ATP1A3 mutations in AHC lie within a cluster in or near transmembrane α-helix TM6, including I810N that is also found in the Myshkin mouse model of AHC. These mutations all substantially reduce Na(+),K(+)-ATPase α3 activity. Herein, we show that Myshkin mice carrying a wild-type Atp1a3 transgene that confers a 16 % increase in brain-specific total Na(+),K(+)-ATPase activity show significant phenotypic improvements compared with non-transgenic Myshkin mice. Interventions to increase the activity of wild-type Na(+),K(+)-ATPase α3 in AHC patients should be investigated further.

  2. Robust ferromagnetism carried by antiferromagnetic domain walls

    NASA Astrophysics Data System (ADS)

    Hirose, Hishiro T.; Yamaura, Jun-Ichi; Hiroi, Zenji

    2017-02-01

    Ferroic materials, such as ferromagnetic or ferroelectric materials, have been utilized as recording media for memory devices. A recent trend for downsizing, however, requires an alternative, because ferroic orders tend to become unstable for miniaturization. The domain wall nanoelectronics is a new developing direction for next-generation devices, in which atomic domain walls, rather than conventional, large domains themselves, are the active elements. Here we show that atomically thin magnetic domain walls generated in the antiferromagnetic insulator Cd2Os2O7 carry unusual ferromagnetic moments perpendicular to the wall as well as electron conductivity: the ferromagnetic moments are easily polarized even by a tiny field of 1 mT at high temperature, while, once cooled down, they are surprisingly robust even in an inverse magnetic field of 7 T. Thus, the magnetic domain walls could serve as a new-type of microscopic, switchable and electrically readable magnetic medium which is potentially important for future applications in the domain wall nanoelectronics.

  3. Robust ferromagnetism carried by antiferromagnetic domain walls

    PubMed Central

    Hirose, Hishiro T.; Yamaura, Jun-ichi; Hiroi, Zenji

    2017-01-01

    Ferroic materials, such as ferromagnetic or ferroelectric materials, have been utilized as recording media for memory devices. A recent trend for downsizing, however, requires an alternative, because ferroic orders tend to become unstable for miniaturization. The domain wall nanoelectronics is a new developing direction for next-generation devices, in which atomic domain walls, rather than conventional, large domains themselves, are the active elements. Here we show that atomically thin magnetic domain walls generated in the antiferromagnetic insulator Cd2Os2O7 carry unusual ferromagnetic moments perpendicular to the wall as well as electron conductivity: the ferromagnetic moments are easily polarized even by a tiny field of 1 mT at high temperature, while, once cooled down, they are surprisingly robust even in an inverse magnetic field of 7 T. Thus, the magnetic domain walls could serve as a new-type of microscopic, switchable and electrically readable magnetic medium which is potentially important for future applications in the domain wall nanoelectronics. PMID:28195565

  4. Robust ferromagnetism carried by antiferromagnetic domain walls.

    PubMed

    Hirose, Hishiro T; Yamaura, Jun-Ichi; Hiroi, Zenji

    2017-02-14

    Ferroic materials, such as ferromagnetic or ferroelectric materials, have been utilized as recording media for memory devices. A recent trend for downsizing, however, requires an alternative, because ferroic orders tend to become unstable for miniaturization. The domain wall nanoelectronics is a new developing direction for next-generation devices, in which atomic domain walls, rather than conventional, large domains themselves, are the active elements. Here we show that atomically thin magnetic domain walls generated in the antiferromagnetic insulator Cd2Os2O7 carry unusual ferromagnetic moments perpendicular to the wall as well as electron conductivity: the ferromagnetic moments are easily polarized even by a tiny field of 1 mT at high temperature, while, once cooled down, they are surprisingly robust even in an inverse magnetic field of 7 T. Thus, the magnetic domain walls could serve as a new-type of microscopic, switchable and electrically readable magnetic medium which is potentially important for future applications in the domain wall nanoelectronics.

  5. Characterization of a Staphylococcal Plasmid Related to pUB110 and Carrying Two Novel Genes, vatC and vgbB, Encoding Resistance to Streptogramins A and B and Similar Antibiotics

    PubMed Central

    Allignet, Jeanine; Liassine, Nadia; El Solh, Névine

    1998-01-01

    We isolated and sequenced a plasmid, named pIP1714 (4,978 bp), which specifies resistance to streptogramins A and B and the mixture of these compounds. pIP1714 was isolated from a Staphylococcus cohnii subsp. cohnii strain found in the environment of a hospital where pristinamycin was extensively used. Resistance to both compounds and related antibiotics is encoded by two novel, probably cotranscribed genes, (i) vatC, encoding a 212-amino-acid (aa) acetyltransferase that inactivates streptogramin A and that exhibits 58.2 to 69.8% aa identity with the Vat, VatB, and SatA proteins, and (ii) vgbB, encoding a 295-aa lactonase that inactivates streptogramin B and that shows 67% aa identity with the Vgb lactonase. pIP1714 includes a 2,985-bp fragment also found in two rolling-circle replication and mobilizable plasmids, pUB110 and pBC16, from gram-positive bacteria. In all three plasmids, the common fragment was delimited by two direct repeats of four nucleotides (GGGC) and included (i) putative genes closely related to repB, which encodes a replication protein, and to pre(mob), which encodes a protein required for conjugative mobilization and site-specific recombination, and (ii) sequences very similar to the double- and single-strand origins (dso, ssoU) and the recombination site, RSA. The antibiotic resistance genes repB and pre(mob) carried by each of these plasmids were found in the same transcriptional orientation. PMID:9661023

  6. Density and composition of an insect population in a field trial of chitinase transgenic and wild-type silver birch (Betula pendula) clones.

    PubMed

    Vihervuori, Liisa; Pasonen, Hanna-Leena; Lyytikäinen-Saarenmaa, Päivi

    2008-12-01

    Fifteen silver birch (Betula pendula Roth) lines carrying a sugar beet chitinase IV gene and eight wild-type birch clones were grown in a field trial. The composition and density of the insect population and the leaf damage caused by insects were monitored and compared between transgenic and wild-type trees. The most abundant insect group in all trees was aphids, and the variation in total insect densities was mainly explained by the variation in aphid densities. Insect densities were generally higher in the transgenic than in the control trees, indicating that the expression of the sugar beet chitinase IV gene had an influence on the suitability of birch leaves to aphids. The level of leaf damage was higher among transgenic than among control trees. Chewing damage was the most common type of leaf damage in all trees. The number of different damage types was higher among the wild-type clones than among the transgenic lines or their controls. The results indicate that the chitinase transgenic trees are more susceptible to aphids and suffer higher levels of leaf damage than the control trees. In the composition of the damage types, the control trees were more similar to the transgenic than to other wild-type trees, indicating that the composition was mostly linked to the genotype of the tree and not to the expression of the transgene. This study provides important information on the ecological interactions of chitinase transgenic trees in the field trial. No clear harmful effects of transgenic chitinase on the biodiversity of insect population were detected.

  7. BLOOD SUBSTITUTES: EVOLUTION FROM NON-CARRYING TO OXYGEN AND GAS CARRYING FLUIDS

    PubMed Central

    Cabrales, Pedro; Intaglietta, Marcos

    2013-01-01

    The development of oxygen (O2) carrying blood substitutes has evolved from the goal of replicating blood O2 transports properties to that of preserving microvascular and organ function, reducing the inherent or potential toxicity of the material used to carry O2, and treating pathologies initiated by anemia and hypoxia. Furthermore, the emphasis has shifted from blood replacement fluid to “O2 therapeutics” that restore tissue oxygenation to specific tissues regions. This review covers the different alternatives, potential and limitations of hemoglobin based O2 carriers (HBOCs) and perfluorocarbon based O2 carriers (PFCOCs), with emphasis on the physiological conditions disturbed in the situation that they will be used. It describes how concepts learned from plasma expanders without O2 carrying capacity can be applied to maintain O2 delivery and summarizes the microvascular responses due to HBOCs and PFCOCs. This review also presents alternative applications of HBOCs and PFCOCs namely: 1) How HBOC O2 affinity can be engineered to target O2 delivery to hypoxic tissues; and 2) How the high gas solubility of PFCOCs provides new opportunities for carrying, dissolving and delivering gases with biological activity. It is concluded that current blood substitutes development has amplified their applications horizon by devising therapeutic functions for oxygen carriers requiring limited O2 delivery capacity restoration. Conversely, full, blood-like O2 carrying capacity re-establishment awaits control of O2 carrier toxicity. PMID:23820271

  8. Global carrying capacity: how many people?

    PubMed

    1992-07-01

    During 1980-85 energy consumption in developing countries increased by 22%, of which 50% was used to maintain current levels of use and 50% pertained to real economic growth. Commercial energy consumption during 1970-89 tripled in developing countries. Population growth alone is expected to increase world energy consumption from the current 13.5 terawatts (13.5 trillion watts) to 18 terawatts by 2025 at the same level of use. The increased level of consumption (4.5 terawatts) is the equivalent of total current commercial energy consumption. One terawatt is equal to energy use from 5 billion barrels of oil yearly, 1 billion tons of coal, or 1.6 billion tons of wood. Economic development will require even greater levels of energy use. Since the oil price increases of the 1970s, developed countries increased their energy consumption by about 33%, even while becoming more fuel efficient. During 1990-2025, if developing countries double their per capita energy use and developed countries reduce their use by 50%, world energy consumption will still be almost 21 terawatts. If consumption remains constant at current levels without any population increase, the oil supply will be exhausted in 40 years. Coal consumption will last hundreds of years but air pollution will worsen, and global warming will be accelerated. Developed countries, which are wealthier, are having difficulty switching to non-fossil fuels, and the prospects for developing countries pose even greater challenges. Slowing growth buys time for technological development. World population is expected to reach 8 billion by 2020. Stabilization of growth at 8 billion would occur only if world fertility averages 1.7 children per woman by 2025. One opinion is that the carrying capacity has been reached with the present population of 5.4 billion. Others say that with changes in consumption and technological developments the earth can sustain 8 billion people. The physical limits are 1) the finite capacity of natural

  9. Transgenic analysis of the thyroid-responsive elements in the alpha-cardiac myosin heavy chain gene promoter.

    PubMed

    Subramaniam, A; Gulick, J; Neumann, J; Knotts, S; Robbins, J

    1993-02-25

    The role of two putative, cis-acting thyroid hormone-responsive elements, TRE1 and TRE2, located at -129 to -149 and -102 to -120, respectively, on the murine alpha-myosin heavy chain (MHC) gene, has been investigated in transgenic mice. These motifs are present in a 4.5-kilobase fragment lying upstream of the transcriptional start site of the mouse alpha-MHC gene: this fragment directs appropriate expression of a reporter gene in transgenic mice (Subramaniam, A., Jones, W. K., Gulick, J., Wert, S., Neumann, J., and Robbins, J. (1991) J. Biol. Chem. 266, 24613-24620). Here, we independently mutate the TRE1 and TRE2 elements by base substitution. The mice were analyzed for transgene expression in different muscle and non-muscle tissues including the atria and ventricles. Normal levels of transgene expression were observed in euthyroid mice carrying a mutation in TRE1. In contrast to these results, mice in which TRE2 was mutated showed reduced levels of CAT activity in both the atria and ventricles, suggesting a previously undefined role for this element in the constitutive up-regulation of the alpha-MHC gene. In hypothyroid mice carrying either of these mutations, the complete cessation of ventricular expression of the chloramphenicol acetyltransferase transcripts that takes place in the alpha-5.5 (wild type) animals did not occur.

  10. [Effects of transgenic crops on soil microorganisms: a review].

    PubMed

    Zhang, Yan-Jun; Xie, Ming; Peng, De-Liang

    2013-09-01

    The worldwide cultivation of transgenic crops not only provides tremendous economic benefits, but also induces the concern about the potential risks of transgenic crops on soil ecosystem in which microorganisms are involved. The potential effects of transgenic crops on soil microorganisms include the direct effects of the transgenic proteins on non-target soil microorganisms, and the indirect effects of the unintentional changes in the chemical compositions of root exudates induced by the introduction of the exogenous transgenic proteins. Most of the studies on transgenic crops suggested that transgenic crops could affect the quantity and structure of soil microbial populations. However, the perceivable effects on the soil microorganisms are inconsistent, with some in significant and others in non-significant, or some with persistent and others with non-persistent. This paper summarized the effects of different transgenic crops on soil microorganisms, and discussed the factors affecting the assessment reliability, including the species of transgenic crops and the experimental technologies and principles. Some issues needed to be paid special attention to in the future studies were put forward.

  11. Auto-reactive B cells in transgenic mice.

    PubMed

    Pasquali, Jean-Louis; Soulas-Sprauel, Pauline; Korganow, Anne-Sophie; Martin, Thierry

    2007-12-01

    In order to understand how the natural occurrence of autoreactive B cells is controlled in normal individuals, and how self reactive B cells can escape this control during diverse clinical situations, many different transgenic mice have been generated expressing self reactive antibodies. In this review, we focus our attention on disease-associated self reactive transgenic models which show the variety of the tolerization mechanisms. The same transgenic lines are also used to analyse the effects of the autoimmune genetic background on the self reactive B cell fate, as well as to study the influence of infectious agents on the behaviour of the auto-reactive transgenic B cells.

  12. [Transferring the Suaeda salsa glutathione S-transferase and catalase genes enhances low temperature stress resistance in transgenic rice seedlings].

    PubMed

    Zhao, Feng-Yun; Wang, Xiao-Yun; Zhao, Yan-Xiu; Zhang, Hui

    2006-04-01

    The GST (glutathione S-transferase) and GST+CAT1 (catalase 1) of Suaeda salsa were introduced into a low temperature-sensitive rice cultivar (Oryza sativa cv. Zhonghua No.11) by Agrobacterium tumefaciens-mediated transformation under the control of cauliflower mosaic virus (CaMV) 35S promoter, and the transformed calli and plantlets were screened on Murashige and Skoog (1962) medium supplemented with hygromycin 25 microg/mL and cefotaxime 300 microg/mL. The putative primary transformants (T(0) generation) were acclimatized at 26 degrees C /22 degrees C in a greenhouse for 7 d, and then transplanted to the field, where they grew up to maturity under outdoor conditions. 25 and 14 independent transgenic lines of T(1) generation carrying the GST and GST+CAT1 genes, respectively, were identified by PCR amplification. Transgene expression was monitored by RNA-blot hybridization using total RNA samples from leaf tissues. To investigate whether expressing the Suaeda salsa GST and GST+CAT1 in transgenic rice increased low temperature stress tolerance, the T(4) 14-day-old transgenic and non-transgenic rice seedlings were transferred to a low temperature (day 7 degrees C/night 4 degrees C) growth chamber for 3-6 d. The experimental data showed that expressing the Suaeda salsa GST and GST+CAT1 enhanced low temperature stress resistance in transgenic rice seedlings. When treated with low temperature, both GST and CAT activity increased in the transformants with the time of temperature treatment. These transgenic rice plant seedlings exhibited a higher level of photosynthetic capacity than those of the non-transgenic control seedlings under low temperature treatment. Whereas, there were lower H(2)O(2) and MDA (malondialdehyde) content, and relative electrolyte leakage through the plasma membrane was also lower in transgenic rice seedlings than in the parent line under low temperature condition. The results also indicated that GST+CAT1 co-expression conferred greater level of low

  13. Potential of MuS1 Transgenic Tobacco for Phytoremediation of the Urban Soils Contaminated with Cadmium

    NASA Astrophysics Data System (ADS)

    Kim, K. H.; Kim, Y. N.; Kim, S. H.

    2010-05-01

    Urban soils are prone to contamination by trace elements such as Cd, Cu, Pb and Zn. Phytoremediation is one of the attractive remediation methods for soils contaminated with trace elements due to its non-destructive and environmentally-friendly characteristic. Scientists have tried to find hyper-accumulator plants in nature or to develop transgenic plant through genetic engineering. This study was carried out to identify a potential of MuS1 transgenic tobacco for phytoremediation of the urban soils contaminated with Cd. MuS1 is known as a multiple stress related gene with several lines. The previous study using RT-PCR showed that the expression of MuS1 gene in tobacco plant induced tolerance to Cd stress. For this study, MuS1 transgenic tobacco and wild-type tobacco (control) were cultivated in a hydroponic system treated with Cd (0, 50, 100 and 200μM Cd) for 3 weeks. At harvest, both tobacco and nutrient solution were collected and were analyzed for Cd. Effect of Cd treatment on morphological change of the tobacco leaves was also observed by variable-pressure scanning electron microscopy (VP-SEM). The tolerance of MuS1 transgenic tobacco to Cd stress was better than that of wild-type tobacco at all Cd levels. Especially, wild-type tobacco showed chlorosis and withering with 200μM Cd treatment, whereas MuS1 transgenic tobacco gradually recovered from Cd damage. Wild-type tobacco accumulated more Cd (4.65mg per plant) than MuS1 transgenic tobacco (2.37mg per plant) with 200μM Cd treatment. Cd translocation rate from root to leaves was 81.8 % for wild-type tobacco compared to 37.1 % for MuS1 transgenic tobacco. Result of VP-SEM showed that the number of trichome in the leaves for wild-type tobacco increased in comparison with that for untreated samples after 3 weeks, while that for MuS1 transgenic tobacco was not changed by Cd treatment. Results showed that the mechanism of the recovery of the MuS1 tobacco plant was not by high level of Cd uptake and accumulation

  14. Transgene detection by digital droplet PCR.

    PubMed

    Moser, Dirk A; Braga, Luca; Raso, Andrea; Zacchigna, Serena; Giacca, Mauro; Simon, Perikles

    2014-01-01

    Somatic gene therapy is a promising tool for the treatment of severe diseases. Because of its abuse potential for performance enhancement in sports, the World Anti-Doping Agency (WADA) included the term 'gene doping' in the official list of banned substances and methods in 2004. Several nested PCR or qPCR-based strategies have been proposed that aim at detecting long-term presence of transgene in blood, but these strategies are hampered by technical limitations. We developed a digital droplet PCR (ddPCR) protocol for Insulin-Like Growth Factor 1 (IGF1) detection and demonstrated its applicability monitoring 6 mice injected into skeletal muscle with AAV9-IGF1 elements and 2 controls over a 33-day period. A duplex ddPCR protocol for simultaneous detection of Insulin-Like Growth Factor 1 (IGF1) and Erythropoietin (EPO) transgenic elements was created. A new DNA extraction procedure with target-orientated usage of restriction enzymes including on-column DNA-digestion was established. In vivo data revealed that IGF1 transgenic elements could be reliably detected for a 33-day period in DNA extracted from whole blood. In vitro data indicated feasibility of IGF1 and EPO detection by duplex ddPCR with high reliability and sensitivity. On-column DNA-digestion allowed for significantly improved target detection in downstream PCR-based approaches. As ddPCR provides absolute quantification, it ensures excellent day-to-day reproducibility. Therefore, we expect this technique to be used in diagnosing and monitoring of viral and bacterial infection, in detecting mutated DNA sequences as well as profiling for the presence of foreign genetic material in elite athletes in the future.

  15. Perspectives on the state of insect transgenics.

    PubMed

    O'Brochta, David A; Handler, Alfred M

    2008-01-01

    Genetic transformation is a critical component to the fundamental genetic analysis of insect species and holds great promise for establishing strains that improve population control and behavior for practical application. This is especially so for insects that are disease vectors, many of which are currently subject to genomic sequence analysis, and intensive population control measures that must be improved for better efficacy and cost-effectiveness. Transposon-mediated germ-line transformation has been the ultimate goal for most fundamental and practical studies, and impressive strides have been made in recent development of transgene vector and marker systems for several mosquito species. This has resulted in rapid advances in functional genomic sequence analysis and new strategies for biological control based on conditional lethality. Importantly, advances have also been made in our ability to use these systems more effectively in terms of enhanced stability and targeting to specific genomic loci. Nevertheless, not all insects are currently amenable to germ-line transformation techniques, and thus advances in transient somatic expression and paratransgenesis have also been critical, if not preferable for some applications. Of particular importance is how this technology will be used for practical application. Early ideas for population replacement of indigenous pests with innocuous transgenic siblings by transposon-vector spread, may require reevaluation in terms of our current knowledge of the behavior of transposons currently available for transformation. The effective implementation of any control program using released transgenics, will also benefit from broadening the perspective of these control measures as being more mainstream than exotic.

  16. Mice carrying a human GLUD2 gene recapitulate aspects of human transcriptome and metabolome development

    PubMed Central

    Li, Qian; Guo, Song; Jiang, Xi; Bryk, Jaroslaw; Naumann, Ronald; Enard, Wolfgang; Tomita, Masaru; Sugimoto, Masahiro; Khaitovich, Philipp; Pääbo, Svante

    2016-01-01

    Whereas all mammals have one glutamate dehydrogenase gene (GLUD1), humans and apes carry an additional gene (GLUD2), which encodes an enzyme with distinct biochemical properties. We inserted a bacterial artificial chromosome containing the human GLUD2 gene into mice and analyzed the resulting changes in the transcriptome and metabolome during postnatal brain development. Effects were most pronounced early postnatally, and predominantly genes involved in neuronal development were affected. Remarkably, the effects in the transgenic mice partially parallel the transcriptome and metabolome differences seen between humans and macaques analyzed. Notably, the introduction of GLUD2 did not affect glutamate levels in mice, consistent with observations in the primates. Instead, the metabolic effects of GLUD2 center on the tricarboxylic acid cycle, suggesting that GLUD2 affects carbon flux during early brain development, possibly supporting lipid biosynthesis. PMID:27118840

  17. Effect of the cauliflower Or transgene on carotenoid accumulation and chromoplast formation in transgenic potato tubers

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Transgenic plants have facilitated our understanding of the functional roles of genes and the metabolic processes affected in plants. Recently, we isolated the Or gene from an orange cauliflower mutant and showed that the Or gene could serve as a novel genetic tool to enrich carotenoid content in tr...

  18. Comparative Proteomics of Leaves from Phytase-Transgenic Maize and Its Non-transgenic Isogenic Variety

    PubMed Central

    Tan, Yanhua; Yi, Xiaoping; Wang, Limin; Peng, Cunzhi; Sun, Yong; Wang, Dan; Zhang, Jiaming; Guo, Anping; Wang, Xuchu

    2016-01-01

    To investigate unintended effects in genetically modified crops (GMCs), a comparative proteomic analysis between the leaves of the phytase-transgenic maize and the non-transgenic plants was performed using two-dimensional gel electrophoresis and mass spectrometry. A total of 57 differentially expressed proteins (DEPs) were successfully identified, which represents 44 unique proteins. Functional classification of the identified proteins showed that these DEPs were predominantly involved in carbohydrate transport and metabolism category, followed by post-translational modification. KEGG pathway analysis revealed that most of the DEPs participated in carbon fixation in photosynthesis. Among them, 15 proteins were found to show protein-protein interactions with each other, and these proteins were mainly participated in glycolysis and carbon fixation. Comparison of the changes in the protein and tanscript levels of the identified proteins showed that most proteins had a similar pattern of changes between proteins and transcripts. Our results suggested that although some significant differences were observed, the proteomic patterns were not substantially different between the leaves of the phytase-transgenic maize and the non-transgenic isogenic type. Moreover, none of the DEPs was identified as a new toxic protein or an allergenic protein. The differences between the leaf proteome might be attributed to both genetic modification and hybrid influence. PMID:27582747

  19. Gene therapy: X-SCID transgene leukaemogenicity.

    PubMed

    Thrasher, Adrian J; Gaspar, H Bobby; Baum, Christopher; Modlich, Ute; Schambach, Axel; Candotti, Fabio; Otsu, Makoto; Sorrentino, Brian; Scobie, Linda; Cameron, Ewan; Blyth, Karen; Neil, Jim; Abina, Salima Hacein-Bey; Cavazzana-Calvo, Marina; Fischer, Alain

    2006-09-21

    Gene therapy has been remarkably effective for the immunological reconstitution of patients with severe combined immune deficiency, but the occurrence of leukaemia in a few patients has stimulated debate about the safety of the procedure and the mechanisms of leukaemogenesis. Woods et al. forced high expression of the corrective therapeutic gene IL2RG, which encodes the gamma-chain of the interleukin-2 receptor, in a mouse model of the disease and found that tumours appeared in a proportion of cases. Here we show that transgenic IL2RG does not necessarily have potent intrinsic oncogenic properties, and argue that the interpretation of this observation with respect to human trials is overstated.

  20. Transgenic Phytoremediation Blasts onto the Scene

    SciTech Connect

    Hooker, Brian S.; Skeen, R S.

    1999-05-01

    The EPA National Priority List contains 22 ammunition production and processing sites that are laden with explosive and propellant wastes. With levels of 2,4,6-trinitrotoluene (TNT) contamination as high as 200 g/kg of solids, some of these sites are literally on the verge of exploding. They also present serious exposure risks to humans and wildlife, as many of these contaminants are also strong toxins and mutagens. In this issue, French et al. describe a new option for cleaning up this dangerous mixture: the use of transgenic plants. They engineered plants to express a bacterial enzyme that can completely denitrify TNT and trinitroglycerin (GTN) into harmless compounds.

  1. Characterization of a Maize Wip1 Promoter in Transgenic Plants

    PubMed Central

    Zhang, Shengxue; Lian, Yun; Liu, Yan; Wang, Xiaoqing; Liu, Yunjun; Wang, Guoying

    2013-01-01

    The Maize Wip1 gene encodes a wound-induced Bowman-Birk inhibitor (BBI) protein which is a type of serine protease inhibitor, and its expression is induced by wounding or infection, conferring resistance against pathogens and pests. In this study, the maize Wip1 promoter was isolated and its function was analyzed. Different truncated Wip1 promoters were fused upstream of the GUS reporter gene and transformed into Arabidopsis, tobacco and rice plants. We found that (1) several truncated maize Wip1 promoters led to strong GUS activities in both transgenic Arabidopsis and tobacco leaves, whereas low GUS activity was detected in transgenic rice leaves; (2) the Wip1 promoter was not wound-induced in transgenic tobacco leaves, but was induced by wounding in transgenic rice leaves; (3) the truncated Wip1 promoter had different activity in different organs of transgenic tobacco plants; (4) the transgenic plant leaves containing different truncated Wip1 promoters had low GUS transcripts, even though high GUS protein level and GUS activities were observed; (5) there was one transcription start site of Wip1 gene in maize and two transcription start sites of GUS in Wip1::GUS transgenic lines; (6) the adjacent 35S promoter which is present in the transformation vectors enhanced the activity of the truncated Wip1 promoters in transgenic tobacco leaves, but did not influence the disability of truncated Wip1231 promoter to respond to wounding signals. We speculate that an ACAAAA hexamer, several CAA trimers and several elements similar to ACAATTAC octamer in the 5′-untranslated region might contribute to the strong GUS activity in Wip1231 transgenic lines, meanwhile, compared to the 5′-untranslated region from Wip1231 transgenic lines, the additional upstream open reading frames (uORFs) in the 5′-untranslated region from Wip1737 transgenic lines might contribute to the lower level of GUS transcript and GUS activity. PMID:24322445

  2. Embryonic fate map of first pharyngeal arch structures in the sox10: kaede zebrafish transgenic model.

    PubMed

    Dougherty, Max; Kamel, George; Shubinets, Valeriy; Hickey, Graham; Grimaldi, Michael; Liao, Eric C

    2012-09-01

    Cranial neural crest cells follow stereotypic patterns of migration to form craniofacial structures. The zebrafish is a powerful vertebrate genetic model where transgenics with reporter proteins under the transcriptional regulation of lineage-specific promoters can be generated. Numerous studies demonstrate that the zebrafish ethmoid plate is embryologically analogous to the mammalian palate. A fate map correlating embryonic cranial neural crest to defined jaw structures would provide a useful context for the morphogenetic analysis of craniofacial development. To that end, the sox10:kaede transgenic was generated, where sox10 provides lineage restriction to the neural crest. Specific regions of neural crest were labeled at the 10-somite stage by photoconversion of the kaede reporter protein. Lineage analysis was carried out during pharyngeal development in wild-type animals, after miR140 injection, and after estradiol treatment. At the 10-somite stage, cranial neural crest cells anterior of the eye contributed to the median ethmoid plate, whereas cells medial to the eye formed the lateral ethmoid plate and trabeculae and a posterior population formed the mandible. miR-140 overexpression and estradiol inhibition of Hedgehog signaling resulted in cleft development, with failed migration of the anterior cell population to form the median ethmoid plate. The sox10:kaede transgenic line provides a useful tool for neural crest lineage analysis. These studies illustrate the advantages of the zebrafish model for application in morphogenetic studies of vertebrate craniofacial development.

  3. Nucleic acid and protein elimination during the sugar manufacturing process of conventional and transgenic sugar beets.

    PubMed

    Klein, J; Altenbuchner, J; Mattes, R

    1998-02-26

    The fate of cellular DNA during the standard purification steps of the sugar manufacturing process from conventional and transgenic sugar beets was determined. Indigenous nucleases of sugar beet cells were found to be active during the first extraction step (raw juice production) which was carried out at 70 degrees C. This and the consecutive steps of the manufacturing process were validated in terms of DNA degradation by competitive PCR of added external DNA. Each step of the process proved to be very efficient in the removal of nucleic acids. Taken together, the purification steps have the potential to reduce the amount of DNA by a factor of > 10(14), exceeding by far the total amount of DNA present in sugar beets. Furthermore, the gene products of the transgenes neomycin phosphotransferase and BNYVV (rhizomania virus) coat protein CP21 were shown to be removed during the purification steps, so that they could not be detected in the resulting white sugar. Thus, sugar obtained from conventional and transgenic beets is indistinguishable or substantially equivalent with respect to purity.

  4. Genotype-Independent Transmission of Transgenic Fluorophore Protein by Boar Spermatozoa

    PubMed Central

    Garrels, Wiebke; Holler, Stephanie; Taylor, Ulrike; Herrmann, Doris; Struckmann, Christina; Klein, Sabine; Barg-Kues, Brigitte; Nowak-Imialek, Monika; Ehling, Christine; Rath, Detlef; Ivics, Zoltán; Niemann, Heiner; Kues, Wilfried A.

    2011-01-01

    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. PMID:22110672

  5. Apoptosis of nur77/N10-transgenic thymocytes involves the Fas/Fas ligand pathway.

    PubMed Central

    Weih, F; Ryseck, R P; Chen, L; Bravo, R

    1996-01-01

    The orphan nuclear receptor Nur77/N10 has recently been demonstrated to be involved in apoptosis of T cell hybridomas. We report here that chronic expression of Nur77/N10 in thymocytes of transgenic mice results in a dramatic reduction of CD4+CD8+ double-positive as well as CD4+CD8- and CD4-CD8+ single-positive cell populations due to an early onset of apoptosis. CD4-CD8- double-negative and CD25+ precursor cells, however, are unaffected. Moreover, nur77/N10-transgenic thymocytes show increased expression of Fas ligand (FasL), while the levels of the Fas receptor (Fas) are not increased. The mouse spontaneous mutant gld (generalized lymphoproliferative disease) carries a point mutation in the extracellular domain of the FasL gene that abolishes the ability of FasL to bind to Fas. Thymuses from nur77/N10-transgenic mice on a gld/gld background have increased cellularity and an almost normal profile of thymocyte subpopulations. Our results demonstrate that one pathway of apoptosis triggered by Nur77/N10 in double-positive thymocytes occurs through the upregulation of FasL expression resulting in increased signaling through Fas. Images Fig. 1 Fig. 2 Fig. 4 PMID:8643610

  6. Characterization of a large human transgene following invasin-mediated delivery in a bacterial artificial chromosome.

    PubMed

    Gillen, Austin E; Lucas, Catherine A; Haussecker, Pei Ling; Kosak, Steven T; Harris, Ann

    2013-10-01

    Bacterial artificial chromosomes (BACs) are widely used in transgenesis, particularly for the humanization of animal models. Moreover, due to their extensive capacity, BACs provide attractive tools to study distal regulatory elements associated with large gene loci. However, despite their widespread use, little is known about the integration dynamics of these large transgenes in mammalian cells. Here, we investigate the post-integration structure of a ~260 kb BAC carrying the cystic fibrosis transmembrane conductance regulator (CFTR) locus following delivery by bacterial invasion and compare this to the outcome of a more routine lipid-based delivery method. We find substantial variability in integrated copy number and expression levels of the BAC CFTR transgene after bacterial invasion-mediated delivery. Furthermore, we frequently observed variation in the representation of different regions of the CFTR transgene within individual cell clones, indicative of BAC fragmentation. Finally, using fluorescence in situ hybridization, we observed that the integrated BAC forms extended megabase-scale structures in some clones that are apparently stably maintained at cell division. These data demonstrate that the utility of large BACs to investigate cis-regulatory elements in the genomic context may be limited by recombination events that complicate their use.

  7. Oct4-enhanced green fluorescent protein transgenic pigs: a new large animal model for reprogramming studies.

    PubMed

    Nowak-Imialek, Monika; Kues, Wilfried A; Petersen, Bjoern; Lucas-Hahn, Andrea; Herrmann, Doris; Haridoss, Srividyameena; Oropeza, Marianne; Lemme, Erika; Schöler, Hans R; Carnwath, Joseph W; Niemann, Heiner

    2011-09-01

    The domesticated pig has emerged as an important tool for development of surgical techniques, advancement of xenotransplantation, creation of important disease models, and preclinical testing of novel cell therapies. However, germ line-competent pluripotent porcine stem cells have not yet been derived. This has been a major obstacle to genetic modification of pigs. The transcription factor Oct4 is essential for the maintenance of pluripotency and for reprogramming somatic cells to a pluripotent state. Here, we report the production of transgenic pigs carrying an 18 kb genomic sequence of the murine Oct4 gene fused to the enhanced green fluorescent protein (EGFP) cDNA (OG2 construct) to allow identification of pluripotent cells by monitoring Oct4 expression by EGFP fluorescence. Eleven viable transgenic piglets were produced by somatic cell nuclear transfer. Expression of the EGFP reporter construct was confined to germ line cells, the inner cell mass and trophectoderm of blastocysts, and testicular germ cells. Reprogramming of fibroblasts from these animals by fusion with pluripotent murine embryonic stem cells or viral transduction with human OCT4, SOX2, KLF4, and c-MYC cDNAs resulted in Oct4-EGFP reactivation. The OG2 pigs have thus proved useful for monitoring reprogramming and the induction and maintenance of pluripotency in porcine cells. In conclusion, the OG2 transgenic pigs are a new large animal model for studying the derivation and maintenance of pluripotent cells, and will be valuable for the development of cell therapy.

  8. Production of early flowering transgenic barley expressing the early flowering allele of Cryptochrome2 gene.

    PubMed

    El-Assal, Salah El-Din; Abd-Alla, Samir M; El-Tarras, Adel A; El-Awady, Mohamed A

    2011-01-01

    This work was carried out in order to develop early flowering barley lines. These lines will be useful to producers by enabling multiple crops within a single season and increasing production. Transgenic barley plants containing the natural early flowering time AtCRY2 allele from the Cape Verde Island (Cvi) ecotype of Arabidopsis have been generated using biolistic transformation. Immature embryo derived calli of two commercially important barley cultivars (El-Dwaser and El-Taif), were transformed using a pCAMBIA-2300 plasmid harboring a genomic fragment containing the AtCRY2-Cvi allele. Transformation was performed utilizing 600 immature embryos for each cultivar. Stable transformation was confirmed in T 0 and T 1 plants by using genomic PCR, RT-PCR and western blot analysis with AtCRY2 specific primers and antibodies, respectively. The transformation efficiency was 5.6% and 3.4% for El-Dwaser and El-Taif cultivars, respectively. Seeds from several T 1 lines were germinated on kanamycin plates and the lines that contained a single locus were selected for further evaluation. The transformed barley plants showed the specific AtCRY2-Cvi flowering phenotype, i.e. early flowering and day length insensitivity, compared to the non transgenic plants. The time to flowering in transgenic T 1 plants was assessed and two lines exhibited flowering more than 25 days earlier than the parental cultivars under short day conditions.

  9. Infertility resulting from transgenic I-PpoI male Anopheles gambiae in large cage trials

    PubMed Central

    Klein, T A; Windbichler, N; Deredec, A; Burt, A; Benedict, M Q

    2012-01-01

    Objectives Anopheles gambiae is the primary vector of malaria in sub-Saharan Africa and is a potential target of genetic control programs. We determined the capacity of male A. gambiae created by germline transformation to introduce infertility into stable age-distribution populations. We also determined effects of the transgenes on life history. Methods Stable age-distribution populations of A. gambiae mosquitoes were established in large indoor cages. Male mosquitoes carrying an I-PpoI homing endonuclease gene were introduced at ×5 and ×10 release rates where they competed with target male mosquitoes for matings. Similar trials were conducted in small cages with an additional ×1 release level. Results Infertility was successfully introduced into all target populations. In supporting experiments, complete female infertility was observed in all strains and species of the A. gambiae complex to which transgenic males were mated. Life history experiments demonstrated that reductions in I-PpoI male vigor exist in the form of reduced adult male emergence, longevity and competitiveness. Discussion A. gambiae I-PpoI males are capable of introducing high levels of infertility in target populations in indoor cage trials. This was accomplished despite losses of vigor resulting from the HEG transgene. These results motivate further trials of sexually I-PpoI A. gambiae in outdoor cage and field trials. PMID:22595271

  10. Overview of expression of hepatitis B surface antigen in transgenic plants.

    PubMed

    Guan, Zheng-jun; Guo, Bin; Huo, Yan-lin; Guan, Zheng-ping; Wei, Ya-hui

    2010-10-28

    Hepatitis B virus (HBV), a pathogen for chronic liver infection, afflicts more than 350 million people world-wide. The effective way to control the virus is to take HBV vaccine. Hepatitis B surface antigen (HBsAg) is an effective protective antigen suitable for vaccine development. At present, "edible" vaccine based on transgenic plants is one of the most promising directions in novel types of vaccines. HBsAg production from transgenic plants has been carried out, and the transgenic plant expression systems have developed from model plants (such as tobacco, potato and tomato) to other various plant platforms. Crude or purified extracts of transformed plants have been found to conduct immunological responses and clinical trials for hepatitis B, which gave the researches of plant-based HBsAg production a big boost. The aim of this review was to summarize the recent data about plant-based HBsAg development including molecular biology of HBsAg gene, selection of expression vector, the expression of HBsAg gene in plants, as well as corresponding immunological responses in animal models or human.

  11. Development of Transgenic Cotton Lines Expressing Allium sativum Agglutinin (ASAL) for Enhanced Resistance against Major Sap-Sucking Pests

    PubMed Central

    Nunna, Hariprasad Rao; Puligundla, Sateesh Kumar; Vudem, Dashavantha Reddy; Khareedu, Venkateswara Rao

    2013-01-01

    Mannose-specific Allium sativum leaf agglutinin encoding gene (ASAL) and herbicide tolerance gene (BAR) were introduced into an elite cotton inbred line (NC-601) employing Agrobacterium-mediated genetic transformation. Cotton transformants were produced from the phosphinothricin (PPT)-resistant shoots obtained after co-cultivation of mature embryos with the Agrobacterium strain EHA105 harbouring recombinant binary vector pCAMBIA3300-ASAL-BAR. PCR and Southern blot analysis confirmed the presence and stable integration of ASAL and BAR genes in various transformants of cotton. Basta leaf-dip assay, northern blot, western blot and ELISA analyses disclosed variable expression of BAR and ASAL transgenes in different transformants. Transgenes, ASAL and BAR, were stably inherited and showed co-segregation in T1 generation in a Mendelian fashion for both PPT tolerance and insect resistance. In planta insect bioassays on T2 and T3 homozygous ASAL-transgenic lines revealed potent entomotoxic effects of ASAL on jassid and whitefly insects, as evidenced by significant decreases in the survival, development and fecundity of the insects when compared to the untransformed controls. Furthermore, the transgenic cotton lines conferred higher levels of resistance (1–2 score) with minimal plant damage against these major sucking pests when bioassays were carried out employing standard screening techniques. The developed transgenics could serve as a potential genetic resource in recombination breeding aimed at improving the pest resistance of cotton. This study represents the first report of its kind dealing with the development of transgenic cotton resistant to two major sap-sucking insects. PMID:24023750

  12. Development of transgenic cotton lines expressing Allium sativum agglutinin (ASAL) for enhanced resistance against major sap-sucking pests.

    PubMed

    Vajhala, Chakravarthy S K; Sadumpati, Vijaya Kumar; Nunna, Hariprasad Rao; Puligundla, Sateesh Kumar; Vudem, Dashavantha Reddy; Khareedu, Venkateswara Rao

    2013-01-01

    Mannose-specific Allium sativum leaf agglutinin encoding gene (ASAL) and herbicide tolerance gene (BAR) were introduced into an elite cotton inbred line (NC-601) employing Agrobacterium-mediated genetic transformation. Cotton transformants were produced from the phosphinothricin (PPT)-resistant shoots obtained after co-cultivation of mature embryos with the Agrobacterium strain EHA105 harbouring recombinant binary vector pCAMBIA3300-ASAL-BAR. PCR and Southern blot analysis confirmed the presence and stable integration of ASAL and BAR genes in various transformants of cotton. Basta leaf-dip assay, northern blot, western blot and ELISA analyses disclosed variable expression of BAR and ASAL transgenes in different transformants. Transgenes, ASAL and BAR, were stably inherited and showed co-segregation in T1 generation in a Mendelian fashion for both PPT tolerance and insect resistance. In planta insect bioassays on T2 and T3 homozygous ASAL-transgenic lines revealed potent entomotoxic effects of ASAL on jassid and whitefly insects, as evidenced by significant decreases in the survival, development and fecundity of the insects when compared to the untransformed controls. Furthermore, the transgenic cotton lines conferred higher levels of resistance (1-2 score) with minimal plant damage against these major sucking pests when bioassays were carried out employing standard screening techniques. The developed transgenics could serve as a potential genetic resource in recombination breeding aimed at improving the pest resistance of cotton. This study represents the first report of its kind dealing with the development of transgenic cotton resistant to two major sap-sucking insects.

  13. Molecular analysis of transgenic melon plants showing virus resistance conferred by direct repeat of movement gene of Cucumber green mottle mosaic virus.

    PubMed

    Ali, Emran Md; Emran, Ali; Tabei, Yutaka; Kobayashi, Kappei; Yamaoka, Naoto; Nishiguchi, Masamichi

    2012-08-01

    Cucumber green mottle mosaic virus (CGMMV) is a major limiting factor in the production of melon plants worldwide. For effective control of this virus using the transgenic approach, the direct repeat of the movement protein gene of CGMMV was used for transforming melon plants by Agrobacterium tumefaciens. PCR and Southern blot analyses of T₃ confirmed that they carried the transgene. Northern blot analysis with total RNA showed that transgene transcript RNA as well as siRNA was observed in all plants tested. Separate leaves or individual plants were inoculated with CGMMV and subjected to ELISA and RNA blot analysis using the coat protein gene probe of the virus. Compared to nontransgenic control, these plants were shown to have high virus resistance. Furthermore, cytosine of the transgene DNA in the plants was methylated. Thus, these results reveal that the transgenic lines were highly resistant to the virus through RNA silencing. Key message High virus resistance was obtained in transgenic melon plants with direct repeat of movement protein gene of Cucumber green mottle mosaic tobamovirus through RNA silencing.

  14. Expression of a potato antimicrobial peptide SN1 increases resistance to take-all pathogen Gaeumannomyces graminis var. tritici in transgenic wheat.

    PubMed

    Rong, Wei; Qi, Lin; Wang, Jingfen; Du, Lipu; Xu, Huijun; Wang, Aiyun; Zhang, Zengyan

    2013-08-01

    Take-all, caused by soil-borne fungus Gaeumannomyces graminis var. tritici (Ggt), is a devastating root disease of wheat (Triticum aestivum) worldwide. Breeding resistant wheat cultivars is the most promising and reliable approach to protect wheat from take-all. Currently, no resistant wheat germplasm is available to breed cultivars using traditional methods. In this study, gene transformation was carried out using Snakin-1 (SN1) gene isolated from potato (Solanum tuberosum) because the peptide shows broad-spectrum antimicrobial activity in vitro. Purified SN1 peptide also inhibits in vitro the growth of Ggt mycelia. By bombardment-mediated method, the gene SN1 was transformed into Chinese wheat cultivar Yangmai 18 to generate SN1 transgenic wheat lines, which were used to assess the effectiveness of the SN1 peptide in protecting wheat from Ggt. Genomic PCR and Southern blot analyses indicated that the alien gene SN1 was integrated into the genomes of five transgenic wheat lines and heritable from T₀ to T₄ progeny. Reverse transcription-PCR and Western blot analyses showed that the introduced SN1 gene was transcribed and highly expressed in the five transgenic wheat lines. Following challenging with Ggt, disease test results showed that compared to segregants lacking the transgene and untransformed wheat plants, these five transgenic wheat lines expressing SN1 displayed significantly enhanced resistance to take-all. These results suggest that SN1 may be a potentially transgenic tool for improving the take-all resistance of wheat.

  15. Production cost analysis and use of pesticides in the transgenic and conventional corn crop [Zea mays (L.)] in the valley of San Juan, Tolima.

    PubMed

    Méndez, Kelly Avila; Chaparro Giraldo, Alejandro; Moreno, Giovanni Reyes; Castro, Carlos Silva

    2011-01-01

    A survey of 10 producers of conventional corn (Hybrids PAC 105 and Maximus) and 10 producers of transgenic corn (Pioneer Hybrid 30T17) was carried out in the municipality of Valle de San Juan in the territorial division of Tolima (Colombia), in order to analyze the differences in production costs and environmental impacts of these two agricultural technologies.  The environmental impacts were determined by calculating the field "Environmental Index Quotient" (EIQ). In the production cost analysis, a difference of 15% was found in benefit of the transgenic technology. The structure of costs of the transgenic technology was benefited by the reduced use of pesticides (insecticides and herbicides). In regards to production, the transgenic technology showed a greater yield, 5.22 ton/ha in comparison to 4.25 ton/ha the conventional technology, thus a 22% difference in yield. Finally, the EIQ calculation showed quantitative differences of 196.12 for the conventional technology (EIQ insecticides 165.14 + EIQ herbicides 30.98), while the transgenic technology was of 4.24 (EIQ insecticides 0 + EIQ herbicides 4.24). These results show a minor environmental impact when using the transgenic technology in comparison to the conventional technology, in regards to the use of insecticides and herbicides in a temporal, spatial and genotypical context analysis. :

  16. Craniosynostosis in transgenic mice overexpressing Nell-1

    PubMed Central

    Zhang, Xinli; Kuroda, Shun’ichi; Carpenter, Dale; Nishimura, Ichiro; Soo, Chia; Moats, Rex; Iida, Keisuke; Wisner, Eric; Hu, Fei-Ya; Miao, Steve; Beanes, Steve; Dang, Catherine; Vastardis, Heleni; Longaker, Michael; Tanizawa, Katsuyuki; Kanayama, Norihiro; Saito, Naoaki; Ting, Kang

    2002-01-01

    Previously, we reported NELL-1 as a novel molecule overexpressed during premature cranial suture closure in patients with craniosynostosis (CS), one of the most common congenital craniofacial deformities. Here we describe the creation and analysis of transgenic mice overexpressing Nell-1. Nell-1 transgenic animals exhibited CS-like phenotypes that ranged from simple to compound synostoses. Histologically, the osteogenic fronts of abnormally closing/closed sutures in these animals revealed calvarial overgrowth and overlap along with increased osteoblast differentiation and reduced cell proliferation. Furthermore, anomalies were restricted to calvarial bone, despite generalized, non-tissue-specific overexpression of Nell-1. In vitro, Nell-1 overexpression accelerated calvarial osteoblast differentiation and mineralization under normal culture conditions. Moreover, Nell-1 overexpression in osteoblasts was sufficient to promote alkaline phosphatase expression and micronodule formation. Conversely, downregulation of Nell-1 inhibited osteoblast differentiation in vitro. In summary, Nell-1 overexpression induced calvarial overgrowth resulting in premature suture closure in a rodent model. Nell-1, therefore, has a novel role in CS development, perhaps as part of a complex chain of events resulting in premature suture closure. On a cellular level, Nell-1 expression may modulate and be both sufficient and required for osteoblast differentiation. PMID:12235118

  17. [Biofuels, food security and transgenic crops].

    PubMed

    Acosta, Orlando; Chaparro-Giraldo, Alejandro

    2009-01-01

    Soaring global food prices are threatening to push more poor people back below the poverty line; this will probably become aggravated by the serious challenge that increasing population and climate changes are posing for food security. There is growing evidence that human activities involving fossil fuel consumption and land use are contributing to greenhouse gas emissions and consequently changing the climate worldwide. The finite nature of fossil fuel reserves is causing concern about energy security and there is a growing interest in the use of renewable energy sources such as biofuels. There is growing concern regarding the fact that biofuels are currently produced from food crops, thereby leading to an undesirable competition for their use as food and feed. Nevertheless, biofuels can be produced from other feedstocks such as lingo-cellulose from perennial grasses, forestry and vegetable waste. Biofuel energy content should not be exceeded by that of the fossil fuel invested in its production to ensure that it is energetically sustainable; however, biofuels must also be economically competitive and environmentally acceptable. Climate change and biofuels are challenging FAO efforts aimed at eradicating hunger worldwide by the next decade. Given that current crops used in biofuel production have not been domesticated for this purpose, transgenic technology can offer an enormous contribution towards improving biofuel crops' environmental and economic performance. The present paper critically presents some relevant relationships between biofuels, food security and transgenic plant technology.

  18. Investigations into the hypothesis of transgenic cannabis.

    PubMed

    Cascini, Fidelia

    2012-05-01

    The unusual concentration of cannabinoids recently found in marijuana samples submitted to the forensic laboratory for chemical analysis prompted an investigation into whether genetic modifications have been made to the DNA of Cannabis sativa L. to increase its potency. Traditional methods for the detection of genetically modified organisms (GMO) were used to analyze herbal cannabis preparations. Our analyses support the hypothesis that marijuana samples submitted to forensic laboratories and characterized by an abnormal level of Δ(9)-THC are the product of breeding selection rather than of transgenic modifications. Further, this research has shown a risk of false positive results associated with the poor quality of the seized samples and probably due to the contamination by other transgenic vegetable products. On the other hand, based on these data, a conclusive distinction between the hypothesis of GMO plant contamination and the other of genetic modification of cannabis cannot be made requiring further studies on comparative chemical and genetic analyses to find out an explanation for the recently detected increased potency of cannabis.

  19. Hydrogen fuel production by transgenic microalgae.

    PubMed

    Melis, Anastasios; Seibert, Michael; Ghirardi, Maria L

    2007-01-01

    This chapter summarizes the state-of-art in the field of green algal H2-production and examines physiological and genetic engineering approaches by which to improve the hydrogen metabolism characteristics of these microalgae. Included in this chapter are emerging topics pertaining to the application of sulfur-nutrient deprivation to attenuate O2-evolution and to promote H2-production, as well as the genetic engineering of sulfate uptake through manipulation of a newly reported sulfate permease in the chloroplast of the model green alga Chlamydomonas reinhardtii. Application of the green algal hydrogenase assembly genes is examined in efforts to confer H2-production capacity to other commercially significant unicellular green algae. Engineering a solution to the O2 sensitivity of the green algal hydrogenase is discussed as an alternative approach to sulfur nutrient deprivation, along with starch accumulation in microalgae for enhanced H2-production. Lastly, current efforts aiming to optimize light utilization in transgenic microalgae for enhanced H2-production under mass culture conditions are presented. It is evident that application of genetic engineering technologies and the use of transgenic green algae will improve prospects for commercial exploitation of these photosynthetic micro-organisms in the generation of H2, a clean and renewable fuel.

  20. L1 integration in a transgenic mouse model

    PubMed Central

    Babushok, Daria V.; Ostertag, Eric M.; Courtney, Christine E.; Choi, Janice M.; Kazazian, Haig H.

    2006-01-01

    To study integration of the human LINE-1 retrotransposon (L1) in vivo, we developed a transgenic mouse model of L1 retrotransposition that displays de novo somatic L1 insertions at a high frequency, occasionally several insertions per mouse. We mapped 3′ integration sites of 51 insertions by Thermal Asymmetric Interlaced PCR (TAIL–PCR). Analysis of integration locations revealed a broad genomic distribution with a modest preference for intergenic regions. We characterized the complete structures of 33 de novo retrotransposition events. Our results highlight the large number of highly truncated L1s, as over 52% (27/51) of total integrants were <1/3 the length of a full-length element. New integrants carry all structural characteristics typical of genomic L1s, including a number with inversions, deletions, and 5′-end microhomologies to the target DNA sequence. Notably, at least 13% (7/51) of all insertions contain a short stretch of extra nucleotides at their 5′ end, which we postulate result from template-jumping by the L1-encoded reverse transcriptase. We propose a unified model of L1 integration that explains all of the characteristic features of L1 retrotransposition, such as 5′ truncations, inversions, extra nucleotide additions, and 5′ boundary and inversion point microhomologies. PMID:16365384

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

    PubMed

    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.

  2. Transgenic Crops and Sustainable Agriculture in the European Context

    ERIC Educational Resources Information Center

    Ponti, Luigi

    2005-01-01

    The rapid adoption of transgenic crops in the United States, Argentina, and Canada stands in strong contrast to the situation in the European Union (EU), where a de facto moratorium has been in place since 1998. This article reviews recent scientific literature relevant to the problematic introduction of transgenic crops in the EU to assess if…

  3. Overview of the investigation of transgenic plums in Romania

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Transgenic plums of Prunus domestica L. transformed with the Plum pox virus coat protein gene (PPV-CP) were the subjects of three experiments undertaken in Romania. In the first experiment, PPV-CP transgenic clones C2, C3, C4, C5, C6 and PT3 were evaluated for Sharka resistance under high natural i...

  4. Overview on the investigations of transgenic plums in Romania

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Transgenic plums of Prunus domestica L. transformed with the Plum pox virus coat protein gene (PPV-CP) were the subjects of three experiments undertaken in Romania. In the first experiment, PPV-CP transgenic clones C2, C3, C4, C5, C6, PT3 and PT5 were evaluated for Sharka resistance under high natu...

  5. Effects of Transgenic Glyphosate-Resistant Crops on Water Quality

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Glyphosate (N-[phosphonomethyl] glycine) is a highly effective, non-selective herbicide. Herbicide-resistant crop (HRC) has been the most successful trait used in transgenic crops throughout the world. Transgenic glyphosate-resistant crops (GRCs) have been commercialized and grown extensively in the...

  6. 2013 North Dakota Transgenic Barley Research and FHB Nursery Report

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Research continues to develop and test new transgenic plants using genes provided by collaborators. As lines are developed in Golden Promise, they are crossed to Conlon for field testing. Transgenic lines developed in Conlon are being crossed to resistant lines developed by the breeding programs. ...

  7. [Application of near-infrared diffuse reflectance spectroscopy to the detection and identification of transgenic corn].

    PubMed

    Rui, Yu-kui; Luo, Yun-bo; Huang, Kun-lun; Wang, Wei-min; Zhang, Lu-da

    2005-10-01

    With the rapid development of the GMO, more and more GMO food has been pouring into the market. Much attention has been paid to GMO labeling under the controversy of GMO safety. Transgenic corns and their parents were scanned by continuous wave of near infrared diffuse reflectance spectroscopy range of 12000-4000 cm(-1); the resolution was 4 cm(-1); scanning was carried out for 64 times; BP algorithm was applied for data processing. The GMO food was easily resolved. Near-infrared diffuse reflectance spectroscopy is unpolluted and inexpensive compared with PCR and ELISA, so it is a very promising detection method for GMO food.

  8. Knock-out and Transgenic Strategies to Improve Neural Transplantation Therapy for Parkinson’s Disease

    DTIC Science & Technology

    2000-07-01

    Parkinson’s disease (PD) are 1) poor survival of grafted fetal neurons and 2) insufficient axonal outgrowth and functional recovery. We carried out experiments aimed at by short-term inhibition by pre-treatment of fetal cells with pharmacological inhibitors of caspases. In our second objective of enhancing axonal growth leading to optimal functional recovery by neuronal transplants, we employed transgenic bcl-2 overexpressing donor cells and similar molecules influencing the growth of axons in the fetal and adult CNS. Use of such molecules could significantly improve

  9. Optimization of Acidothermus Celluloyticus Endoglucanase (E1) Production in Transgenic Tobacco Plants by Transcriptional, Post-transcription and Post-Translational Modification

    SciTech Connect

    Dai, Ziyu; Hooker, Brian S.; Quesenberry, Ryan D.; Thomas, S. R.

    2005-10-01

    Biochemical characteristics of Acidothermus cellulolyticus endoglucanase (E1) and its physiological effects in transgenic tobacco (Nicotiana tabacum) has been studied previously. In an attempt to obtain a high level of production of intact E1 in transgenic plants, the E1 gene was expressed under the control of strong Mac promoter (a hybrid promoter of manopine synthase promoter and cauliflower mosaic virus 35S promoter enhancer region) or tomato Rubisco small subunit (RbcS-3C) promoter with different 5’ untranslated leader (UTL) sequence and targeted to different subcellular comartmentations with various transit peptides. The expression of E1 protein in transgenic tobacco plants was determined via E1 activity, protein immunobloting, and RNA gel-blotting analyses. Effects of different transit peptides on E1 protein production and its stability were examined in transgenic tobacco plants carrying one of six transgene expression vectors with the same (Mac) promoter and transcription terminator (Tmas). Transgenic tobacco plants with apoplast transit peptide (Mm-apo) had the highest average E1 activity and protein accumulation , while E1 protein was more stable in transgenic plants with no transit peptide (Mm) than others. The E1 expression under tomato RbcS-3C promoter was higher than that under Mac promoter based on the average E1 activity, E1 protein accumulation, and RNA gel-blotting. The E1 expression was increased more than two fold when the 5’-UTL of alfalfa mosaic virus RNA4 gene replaced the UTL of RbcS-3C promoter, while the UTL of alfalfa mosaic virus RNA4 gene was less effective than the UTL of Mac promoter. The optimal combination of promoter, 5’-UTL, and subcellular compartmentation (transit peptide) for E1 protein production in transgenic tobacco plants are discussed.

  10. Small Interfering RNAs That Trigger Posttranscriptional Gene Silencing Are Not Required for the Histone H3 Lys9 Methylation Necessary for Transgenic Tandem Repeat Stabilization in Neurospora crassa†

    PubMed Central

    Chicas, Agustin; Forrest, Emma C.; Sepich, Silvia; Cogoni, Carlo; Macino, Giuseppe

    2005-01-01

    In Neurospora crassa, the introduction of a transgene can lead to small interfering RNA (siRNA)-mediated posttranscriptional gene silencing (PTGS) of homologous genes. siRNAs can also guide locus-specific methylation of Lys9 of histone H3 (Lys9H3) in Schizosaccharomyces pombe. Here we tested the hypothesis that transgenically derived siRNAs may contemporaneously both activate the PTGS mechanism and induce chromatin modifications at the transgene and the homologous endogenous gene. We carried out chromatin immunoprecipitation using a previously characterized albino-1 (al-1) silenced strain but detected no alterations in the pattern of histone modifications at the endogenous al-1 locus, suggesting that siRNAs produced from the transgenic locus do not trigger modifications in trans of those histones tested. Instead, we found that the transgenic locus was hypermethylated at Lys9H3 in our silenced strain and remained hypermethylated in the quelling defective mutants (qde), further demonstrating that the PTGS machinery is dispensable for Lys9H3 methylation. However, we found that a mutant in the histone Lys9H3 methyltransferase dim-5 was unable to maintain PTGS, with transgenic copies being rapidly lost, resulting in reversion of the silenced phenotype. These results indicate that the defect in PTGS of the Δdim-5 strain is due to the inability to maintain the transgene in tandem, suggesting a role for DIM-5 in stabilizing such repeated sequences. We conclude that in Neurospora, siRNAs produced from the transgenic locus are used in the RNA-induced silencing complex-mediated PTGS pathway and do not communicate with an RNAi-induced initiation of transcriptional gene silencing complex to effect chromatin-based silencing. PMID:15831483

  11. Small interfering RNAs that trigger posttranscriptional gene silencing are not required for the histone H3 Lys9 methylation necessary for transgenic tandem repeat stabilization in Neurospora crassa.

    PubMed

    Chicas, Agustin; Forrest, Emma C; Sepich, Silvia; Cogoni, Carlo; Macino, Giuseppe

    2005-05-01

    In Neurospora crassa, the introduction of a transgene can lead to small interfering RNA (siRNA)-mediated posttranscriptional gene silencing (PTGS) of homologous genes. siRNAs can also guide locus-specific methylation of Lys9 of histone H3 (Lys9H3) in Schizosaccharomyces pombe. Here we tested the hypothesis that transgenically derived siRNAs may contemporaneously both activate the PTGS mechanism and induce chromatin modifications at the transgene and the homologous endogenous gene. We carried out chromatin immunoprecipitation using a previously characterized albino-1 (al-1) silenced strain but detected no alterations in the pattern of histone modifications at the endogenous al-1 locus, suggesting that siRNAs produced from the transgenic locus do not trigger modifications in trans of those histones tested. Instead, we found that the transgenic locus was hypermethylated at Lys9H3 in our silenced strain and remained hypermethylated in the quelling defective mutants (qde), further demonstrating that the PTGS machinery is dispensable for Lys9H3 methylation. However, we found that a mutant in the histone Lys9H3 methyltransferase dim-5 was unable to maintain PTGS, with transgenic copies being rapidly lost, resulting in reversion of the silenced phenotype. These results indicate that the defect in PTGS of the Deltadim-5 strain is due to the inability to maintain the transgene in tandem, suggesting a role for DIM-5 in stabilizing such repeated sequences. We conclude that in Neurospora, siRNAs produced from the transgenic locus are used in the RNA-induced silencing complex-mediated PTGS pathway and do not communicate with an RNAi-induced initiation of transcriptional gene silencing complex to effect chromatin-based silencing.

  12. Advancing environmental risk assessment for transgenic biofeedstock crops.

    PubMed

    Wolt, Jeffrey D

    2009-11-02

    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.

  13. Advancing environmental risk assessment for transgenic biofeedstock crops

    PubMed Central

    Wolt, Jeffrey D

    2009-01-01

    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. PMID:19883509

  14. Use of transgenic Aedes aegypti in Brazil: risk perception and assessment

    PubMed Central

    Aragão, Francisco José Lima; Colli, Walter; Dellagostin, Odir Antônio; Finardi-Filho, Flávio; Hirata, Mario Hiroyuki; Lira-Neto, Amaro de Castro; Almeida de Melo, Marcia; Nepomuceno, Alexandre Lima; Gorgônio da Nóbrega, Francisco; Delfino de Sousa, Gutemberg; Valicente, Fernando Hercos; Zanettini, Maria Helena Bodanese

    2016-01-01

    Abstract The OX513A strain of Aedes aegypti, which was developed by the British company Oxitec, expresses a self-limiting transgene that prevents larvae from developing to adulthood. In April 2014, the Brazilian National Technical Commission on Biosafety completed a risk assessment of OX513A and concluded that the strain did not present new biological risks to humans or the environment and could be released in Brazil. At that point, Brazil became the first country to approve the unconstrained release of a genetically modified mosquito. During the assessment, the commission produced a comprehensive list of – and systematically analysed – the perceived hazards. Such hazards included the potential survival to adulthood of immature stages carrying the transgene – should the transgene fail to be expressed or be turned off by exposure to sufficient environmental tetracycline. Other perceived hazards included the potential allergenicity and/or toxicity of the proteins expressed by the gene, the potential for gene flow or increased transmission of human pathogens and the occupation of vacant breeding sites by other vector species. The Zika epidemic both elevated the perceived importance of Ae. aegypti as a vector – among policy-makers and regulators as well as the general public – and increased concerns over the release of males of the OX513A strain. We have therefore reassessed the potential hazards. We found that release of the transgenic mosquitoes would still be both safe and of great potential value in the control of diseases spread by Ae. aegypti, such as chikungunya, dengue and Zika. PMID:27843167

  15. Progressive squamous epithelial neoplasia in K14-human papillomavirus type 16 transgenic mice.

    PubMed Central

    Arbeit, J M; Münger, K; Howley, P M; Hanahan, D

    1994-01-01

    To model human papillomavirus-induced neoplastic progression, expression of the early region of human papillomavirus type 16 (HPV16) was targeted to the basal cells of the squamous epithelium in transgenic mice, using a human keratin 14 (K14) enhancer/promoter. Twenty-one transgenic founder mice were produced, and eight lines carrying either wild-type or mutant HPV16 early regions that did not express the E1 or E2 genes were established. As is characteristic of human cancers, the E6 and E7 genes remained intact in these mutants. The absence of E1 or E2 function did not influence the severity of the phenotype that eventually developed in the transgenic mice. Hyperplasia, papillomatosis, and dysplasia appeared at multiple epidermal and squamous mucosal sites, including ear and truncal skin, face, snout and eyelids, and anus. The ears were the most consistently affected site, with pathology being present in all lines with 100% penetrance. This phenotype also progressed through discernible stages. An initial mild hyperplasia was followed by hyperplasia, which further progressed to dysplasia and papillomatosis. During histopathological progression, there was an incremental increase in cellular DNA synthesis, determined by 5-bromo-2'-deoxyuridine incorporation, and a profound perturbation in keratinocyte terminal differentiation, as revealed by immunohistochemistry to K5, K14, and K10 and filaggrin. These K14-HPV16 transgenic mice present an opportunity to study the role of the HPV16 oncogenes in the neoplastic progression of squamous epithelium and provide a model with which to identify genetic and epigenetic factors necessary for carcinogenesis. Images PMID:7515971

  16. GATA4 transgenic mice as an in vivo model of congenital heart disease

    PubMed Central

    HAN, HUA; CHEN, YU; LIU, GANG; HAN, ZENGQIANG; ZHAO, ZHOU; TANG, YIN

    2015-01-01

    Our previous study indicated that 8 patients from a family with a history of congenital heart disease had simple atrial septal defect (ASD) and carried the same mutation at codon 310 in the GATA4 gene. In the present study, to identify the functional defects caused by this mutation in an in vivo model, the transgene DNA constructs were microinjected into mice to generate a transgenic mouse model. The mice were genotyped using PCR and DNA sequencing. Protein expression was measured by western blot analysis. qPCR was used to determine the copy number of the transgenes. The heart tissue was fixed and sectioned by conventional procedures. The Vevo 2000 system was used to perform echocardiography on the mice. The expression of GATA4 target genes was measured using the real-time PCR system. The incidence of ASD in the heterozygous transgenic mice was found to be greater than that in the wild-type control mice (P<0.05). In addition, the expression of α-myosin heavy chain (α-MHC) in the heart tissues from the homozygous mice was lower than that in the heart tissues from their wild-type littermates (P<0.05). In conclusion, these results suggest that the introduction of GATA4 M310V negatively affects the normal expression of α-MHC. In accordance with previous findings on GATA4 mutation screening and in vitro experiments, this study confirms that GATA4 M310V mutation may lead to the development of the congenital heart defect, ASD. PMID:25873328

  17. Recloned dogs derived from adipose stem cells of a transgenic cloned beagle.

    PubMed

    Oh, Hyun Ju; Park, Jung Eun; Kim, Min Jung; Hong, So Gun; Ra, Jeong Chan; Jo, Jung Youn; Kang, Sung Keun; Jang, Goo; Lee, Byeong Chun

    2011-04-15

    A number of studies have postulated that efficiency in mammalian cloning is inversely correlated with donor cell differentiation status and may be increased by using undifferentiated cells as nuclear donors. Here, we attempted the recloning of dogs by nuclear transfer of canine adipose tissue-derived mesenchymal stem cells (cAd-MSCs) from a transgenic cloned beagle to determine if cAd-MSCs can be a suitable donor cell type. In order to isolate cAd-MSCs, adipose tissues were collected from a transgenic cloned beagle produced by somatic cell nuclear transfer (SCNT) of canine fetal fibroblasts modified genetically with a red fluorescent protein (RFP) gene. The cAd-MSCs expressed the RFP gene and cell-surface marker characteristics of MSCs including CD29, CD44 and thy1.1. Furthermore, cAd-MSCs underwent osteogenic, adipogenic, myogenic, neurogenic and chondrogenic differentiation when exposed to specific differentiation-inducing conditions. In order to investigate the developmental potential of cAd-MSCs, we carried out SCNT. Fused-couplets (82/109, 75.2%) were chemically activated and transferred into the uterine tube of five naturally estrus-synchronized surrogates. One of them (20%) maintained pregnancy and subsequently gave birth to two healthy cloned pups. The present study demonstrated for the first time the successful production of cloned beagles by nuclear transfer of cAd-MSCs. Another important outcome of the present study is the successful recloning of RFP-expressing transgenic cloned beagle pups by nuclear transfer of cells derived from a transgenic cloned beagle. In conclusion, the present study demonstrates that adipose stem cells can be a good nuclear donor source for dog cloning.

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

    PubMed

    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.

  19. Minute Pirate Bug (Orius Insidiosus Say) populations on transgenic and non-transgenic maize using different sampling techniques

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Field experiments were conducted to evaluate the populations of minute pirate bug [Orius insidiosus (Say)] using visual, sticky cards, and destructive sampling techniques in transgenic and non-transgenic maize in three locations in Nebraska (Mead, Clay Center, and Concord), United States of America,...

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

    PubMed

    Pradhan, Bhola Shankar; Majumdar, Subeer S

    2016-03-08

    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.

  1. Estrogen and progesterone receptors have distinct roles in the establishment of the hyperplastic phenotype in PR-A transgenic mice

    SciTech Connect

    Simian, Marina; Bissell, Mina J.; Barcellos-Hoff, Mary Helen; Shyamala, Gopalan

    2009-05-11

    Expression of the A and B forms of progesterone receptor (PR) in an appropriate ratio is critical for mammary development. Mammary glands of PR-A transgenic mice, carrying an additional A form of PR as a transgene, exhibit morphological features associated with the development of mammary tumors. Our objective was to determine the roles of estrogen (E) and progesterone (P) in the genesis of mammary hyperplasias/preneoplasias in PR-A transgenics. We subjected PR-A mice to hormonal treatments and analyzed mammary glands for the presence of hyperplasias and used BrdU incorporation to measure proliferation. Quantitative image analysis was carried out to compare levels of latency-associated peptide and transforming growth factor beta 1 (TGF{beta}1) between PR-A and PR-B transgenics. Basement membrane disruption was examined by immunofluorescence and proteolytic activity by zymography. The hyperplastic phenotype of PR-A transgenics is inhibited by ovariectomy, and is reversed by treatment with E + P. Studies using the antiestrogen ICI 182,780 or antiprogestins RU486 or ZK 98,299 show that the increase in proliferation requires signaling through E/estrogen receptor alpha but is not sufficient to give rise to hyperplasias, whereas signaling through P/PR has little impact on proliferation but is essential for the manifestation of hyperplasias. Increased proliferation is correlated with decreased TGF{beta}1 activation in the PR-A transgenics. Analysis of basement membrane integrity showed loss of laminin-5, collagen III and collagen IV in mammary glands of PR-A mice, which is restored by ovariectomy. Examination of matrix metalloproteases (MMPs) showed that total levels of MMP-2 correlate with the steady-state levels of PR, and that areas of laminin-5 loss coincide with those of activation of MMP-2 in PR-A transgenics. Activation of MMP-2 is dependent on treatment with E and P in ovariectomized wild-type mice, but is achieved only by treatment with P in PR-A mice. These data

  2. WP1: transgenic opto-animals

    NASA Astrophysics Data System (ADS)

    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.

  3. T cell immunity using transgenic B lymphocytes

    NASA Astrophysics Data System (ADS)

    Gerloni, Mara; Rizzi, Marta; Castiglioni, Paola; Zanetti, Maurizio

    2004-03-01

    Adaptive immunity exists in all vertebrates and plays a defense role against microbial pathogens and tumors. T cell responses begin when precursor T cells recognize antigen on specialized antigen-presenting cells and differentiate into effector cells. Currently, dendritic cells are considered the only cells capable of stimulating T lymphocytes. Here, we show that mature naïve B lymphocytes can be genetically programmed by using nonviral DNA and turned into powerful antigen-presenting cells with a dual capacity of synthesis and presentation of antigen to T cells in vivo. A single i.v. injection of transgenic lymphocytes activates T cell responses reproducibly and specifically even at very low cell doses (102). We also demonstrate that T cell priming can occur in the absence of dendritic cells and results in immunological memory with protective effector functions. These findings disclose aspects in the regulation of adaptive immunity and indicate possibilities for vaccination against viruses and cancer in humans.

  4. Viral vectors: from virology to transgene expression

    PubMed Central

    Bouard, D; Alazard-Dany, N; Cosset, F-L

    2009-01-01

    In the late 1970s, it was predicted that gene therapy would be applied to humans within a decade. However, despite some success, gene therapy has still not become a routine practise in medicine. In this review, we will examine the problems, both experimental and clinical, associated with the use of viral material for transgenic insertion. We shall also discuss the development of viral vectors involving the most important vector types derived from retroviruses, adenoviruses, herpes simplex viruses and adeno-associated viruses. This article is part of a themed section on Vector Design and Drug Delivery. For a list of all articles in this section see the end of this paper, or visit: http://www3.interscience.wiley.com/journal/121548564/issueyear?year=2009 PMID:18776913

  5. Novel transgenic rice-based vaccines.

    PubMed

    Azegami, Tatsuhiko; Itoh, Hiroshi; Kiyono, Hiroshi; Yuki, Yoshikazu

    2015-04-01

    Oral vaccination can induce both systemic and mucosal antigen-specific immune responses. To control rampant mucosal infectious diseases, the development of new effective oral vaccines is needed. Plant-based vaccines are new candidates for oral vaccines, and have some advantages over the traditional vaccines in cost, safety, and scalability. Rice seeds are attractive for vaccine production because of their stability and resistance to digestion in the stomach. The efficacy of some rice-based vaccines for infectious, autoimmune, and other diseases has been already demonstrated in animal models. We reported the efficacy in mice, safety, and stability of a rice-based cholera toxin B subunit vaccine called MucoRice-CTB. To advance MucoRice-CTB for use in humans, we also examined its efficacy and safety in primates. The potential of transgenic rice production as a new mucosal vaccine delivery system is reviewed from the perspective of future development of effective oral vaccines.

  6. Transgenic maize plants by tissue electroporation.

    PubMed Central

    D'Halluin, K; Bonne, E; Bossut, M; De Beuckeleer, M; Leemans, J

    1992-01-01

    In this paper, we describe the transformation of regenerable maize tissues by electroporation. In many maize lines, immature zygotic embryos can give rise to embryogenic callus cultures from which plants can be regenerated. Immature zygotic embryos or embryogenic type I calli were wounded either enzymatically or mechanically and subsequently electroporated with a chimeric gene encoding neomycin phosphotransferase (neo). Transformed embryogenic calli were selected from electroporated tissues on kanamycin-containing media and fertile transgenic maize plants were regenerated. The neo gene was transmitted to the progeny of kanamycin-resistant transformants in a Mendelian fashion. This showed that all transformants were nonchimeric, suggesting that transformation and regeneration are a single-cell event. The maize transformation procedure presented here does not require the establishment of genotype-dependent embryogenic type II callus or cell suspension cultures and facilitates the engineering of new traits into agronomically relevant maize inbred lines. PMID:1334743

  7. Magnetic biomineralisation in Huntington's disease transgenic mice

    NASA Astrophysics Data System (ADS)

    Beyhum, W.; Hautot, D.; Dobson, J.; Pankhurst, Q. A.

    2005-01-01

    The concentration levels of biogenic magnetite nanoparticles in transgenic R6/2 Huntington's disease (HD) mice have been investigated, using seven control and seven HD mice each from an 8 week-old litter and from a 12 week-old litter. Hysteresis and isothermal remnant magnetisation data were collected on a SQUID magnetometer, and analysed using a model comprising dia/paramagnetic, ferrimagnetic and superparamagnetic contributions, to extract the magnetite and ferritin concentrations present. It was found that magnetite was present in both superparamagnetic and blocked states. A larger spread and higher concentration of magnetite levels was found in the diseased mice for both the 8 week-old and 12 week-old batches, compared to the controls.

  8. A Transgenic Mouse Model of Poliomyelitis.

    PubMed

    Koike, Satoshi; Nagata, Noriyo

    2016-01-01

    Transgenic mice (tg mice) that express the human poliovirus receptor (PVR), CD155, are susceptible to poliovirus and develop a neurological disease that resembles human poliomyelitis. Assessment of the neurovirulence levels of poliovirus strains, including mutant viruses produced by reverse genetics, circulating vaccine-derived poliovirus, and vaccine candidates, is useful for basic research of poliovirus pathogenicity, the surveillance of circulating polioviruses, and the quality control of oral live poliovirus vaccines, and does not require the use of monkeys. Furthermore, PVR-tg mice are useful for studying poliovirus tissue tropism and host immune responses. PVR-tg mice can be bred with mice deficient in the genes involved in viral pathogenicity. This report describes the methods used to analyze the pathogenicity and immune responses of poliovirus using the PVR-tg mouse model.

  9. High-value products from transgenic maize.

    PubMed

    Naqvi, Shaista; Ramessar, Koreen; Farré, Gemma; Sabalza, Maite; Miralpeix, Bruna; Twyman, Richard M; Capell, Teresa; Zhu, Changfu; Christou, Paul

    2011-01-01

    Maize (also known as corn) is a domesticated cereal grain that has been grown as food and animal feed for tens of thousands of years. It is currently the most widely grown crop in the world, and is used not only for food/feed but also to produce ethanol, industrial starches and oils. Maize is now at the beginning of a new agricultural revolution, where the grains are used as factories to synthesize high-value molecules. In this article we look at the diversity of high-value products from maize, recent technological advances in the field and the emerging regulatory framework that governs how transgenic maize plants and their products are grown, used and traded.

  10. Inheritance of transgenes in transgenic Bt lines resistance to Helicoerpa armigera in upland cotton.

    PubMed

    Zhang, Baolong; Guo, Wangzhen; Zhang, Tianzhen

    2013-01-01

    Six transgenic Bt cotton cultivars (lines) including GKsu12, GK19, MR1, GK5, 109B, and SGK1 are highly resistant to bollworm from the seedling to boll-setting stages in bioassays with detached cotton leaves, though there are differences in resistant level and Bt toxin content in these transgenic cottons. Genetics analysis reveals that the resistance to Helicoverpa armigera in these six transgenic Bt cotton cultivars (lines) are controlled by one pair of dominant genes. Allelic tests further demonstrate some populations are in Mendel segregation for two nonallelic genes, i.e., the inserted Bt gene in GKsu12 is nonallelic to that of SGK1, GK5, 109B, and GK19 and Bt genes in GK19 and SGK1 are likely inserted in the same or in close proximity (genetically closely linked), while some F(2) produce abnormal segregation patterns, with a segregation of resistance to Helicoerpa armigera vary between 15:1 and 3:1, though their Bt segregation fit into 15:1 by PCR analysis, suggesting Bt gene silence in these populations. Two genes silence may occur in these populations due to the homologous sequence by crossing since the silenced individuals accounted for 1/16 of the F(2) populations for allelic test. To those silenced populations, one of their parents all showed high resistance to bollworm.

  11. Transgenic Studies with a Keratin Promoter-Driven Growth Hormone Transgene: Prospects for Gene Therapy

    NASA Astrophysics Data System (ADS)

    Wang, Xiaoming; Zinkel, Sandra; Polonsky, Kenneth; Fuchs, Elaine

    1997-01-01

    Keratinocytes are potentially appealing vehicles for the delivery of secreted gene products because they can be transferred to human skin by the relatively simple procedure of grafting. Adult human keratinocytes can be efficiently propagated in culture with sufficient proliferative capacity to produce enough epidermis to cover the body surface of an average adult. However, the feasibility of delivering secreted proteins through skin grafting rests upon (i) the strength of the promoter in keratinocytes and (ii) the efficiency of protein transport through the basement membrane of the stratified epithelium and into the bloodstream. In this paper, we use transgenic technology to demonstrate that the activity of the human keratin 14 promoter remains high in adult skin and that keratinocyte-derived human growth hormone (hGH) can be produced, secreted, and transported to the bloodstream of mice with efficiency that is sufficient to exceed by an order of magnitude the circulating hGH concentration in growing children. Transgenic skin grafts from these adults continue to produce and secrete hGH stably, at ≈ 1/10 physiological levels in the bloodstream of nontransgenic recipient mice. These studies underscore the utility of the keratin 14 promoter for expressing foreign transgenes in keratinocytes and demonstrate that keratinocytes can be used as effective vehicles for transporting factors to the bloodstream and for eliciting metabolic changes. These findings have important implications for considering the keratinocyte as a possible vehicle for gene therapy.

  12. Human antibody production in transgenic animals.

    PubMed

    Brüggemann, Marianne; Osborn, Michael J; Ma, Biao; Hayre, Jasvinder; Avis, Suzanne; Lundstrom, Brian; Buelow, Roland

    2015-04-01

    Fully human antibodies from transgenic animals account for an increasing number of new therapeutics. After immunization, diverse human monoclonal antibodies of high affinity can be obtained from transgenic rodents, while large animals, such as transchromosomic cattle, have produced respectable amounts of specific human immunoglobulin (Ig) in serum. Several strategies to derive animals expressing human antibody repertoires have been successful. In rodents, gene loci on bacterial artificial chromosomes or yeast artificial chromosomes were integrated by oocyte microinjection or transfection of embryonic stem (ES) cells, while ruminants were derived from manipulated fibroblasts with integrated human chromosome fragments or human artificial chromosomes. In all strains, the endogenous Ig loci have been silenced by gene targeting, either in ES or fibroblast cells, or by zinc finger technology via DNA microinjection; this was essential for optimal production. However, comparisons showed that fully human antibodies were not as efficiently produced as wild-type Ig. This suboptimal performance, with respect to immune response and antibody yield, was attributed to imperfect interaction of the human constant region with endogenous signaling components such as the Igα/β in mouse, rat or cattle. Significant improvements were obtained when the human V-region genes were linked to the endogenous CH-region, either on large constructs or, separately, by site-specific integration, which could also silence the endogenous Ig locus by gene replacement or inversion. In animals with knocked-out endogenous Ig loci and integrated large IgH loci, containing many human Vs, all D and all J segments linked to endogenous C genes, highly diverse human antibody production similar to normal animals was obtained.

  13. An evaluation of new and established methods to determine T‐DNA copy number and homozygosity in transgenic plants.

    PubMed Central

    Głowacka, Katarzyna; Kromdijk, Johannes; Leonelli, Lauriebeth; Niyogi, Krishna K.; Clemente, Tom E.

    2016-01-01

    Abstract Stable transformation of plants is a powerful tool for hypothesis testing. A rapid and reliable evaluation method of the transgenic allele for copy number and homozygosity is vital in analysing these transformations. Here the suitability of Southern blot analysis, thermal asymmetric interlaced (TAIL‐)PCR, quantitative (q)PCR and digital droplet (dd)PCR to estimate T‐DNA copy number, locus complexity and homozygosity were compared in transgenic tobacco. Southern blot analysis and ddPCR on three generations of transgenic offspring with contrasting zygosity and copy number were entirely consistent, whereas TAIL‐PCR often underestimated copy number. qPCR deviated considerably from the Southern blot results and had lower precision and higher variability than ddPCR. Comparison of segregation analyses and ddPCR of T1 progeny from 26 T0 plants showed that at least 19% of the lines carried multiple T‐DNA insertions per locus, which can lead to unstable transgene expression. Segregation analyses failed to detect these multiple copies, presumably because of their close linkage. This shows the importance of routine T‐DNA copy number estimation. Based on our results, ddPCR is the most suitable method, because it is as reliable as Southern blot analysis yet much faster. A protocol for this application of ddPCR to large plant genomes is provided. PMID:26670088

  14. Successful recovery of transgenic cowpea (Vigna unguiculata) using the 6-phosphomannose isomerase gene as the selectable marker.

    PubMed

    Bakshi, Souvika; Saha, Bedabrata; Roy, Nand Kishor; Mishra, Sagarika; Panda, Sanjib Kumar; Sahoo, Lingaraj

    2012-06-01

    A new method for obtaining transgenic cowpea was developed using positive selection based on the Escherichia coli 6-phosphomannose isomerase gene as the selectable marker and mannose as the selective agent. Only transformed cells were capable of utilizing mannose as a carbon source. Cotyledonary node explants from 4-day-old in vitro-germinated seedlings of cultivar Pusa Komal were inoculated with Agrobacterium tumefaciens strain EHA105 carrying the vector pNOV2819. Regenerating transformed shoots were selected on medium supplemented with a combination of 20 g/l mannose and 5 g/l sucrose as carbon source. The transformed shoots were rooted on medium devoid of mannose. Transformation efficiency based on PCR analysis of individual putative transformed shoots was 3.6%. Southern blot analysis on five randomly chosen PCR-positive plants confirmed the integration of the pmi transgene. Qualitative reverse transcription (qRT-PCR) analysis demonstrated the expression of pmi in T₀ transgenic plants. Chlorophenol red (CPR) assays confirmed the activity of PMI in transgenic plants, and the gene was transmitted to progeny in a Mendelian fashion. The transformation method presented here for cowpea using mannose selection is efficient and reproducible, and could be used to introduce a desirable gene(s) into cowpea for biotic and abiotic stress tolerance.

  15. Development of Transgenic Mice Containing an Introduced Stop Codon on the Human Methylmalonyl-CoA Mutase Locus

    PubMed Central

    Buck, Nicole E.; Dashnow, Harriet; Pitt, James J.; Wood, Leonie R.; Peters, Heidi L.

    2012-01-01

    The mutation R403stop was found in an individual with mut0 methylmalonic aciduria (MMA) which resulted from a single base change of C→T in exon 6 of the methylmalonyl-CoA mutase gene (producing a TGA stop codon). In order to accurately model the human MMA disorder we introduced this mutation onto the human methylmalonyl-CoA mutase locus of a bacterial artificial chromosome. A mouse model was developed using this construct. The transgene was found to be intact in the mouse model, with 7 copies integrated at a single site in chromosome 3. The phenotype of the hemizygous mouse was unchanged until crossed against a methylmalonyl-CoA mutase knockout mouse. Pups with no endogenous mouse methylmalonyl-CoA mutase and one copy of the transgene became ill and died within 24 hours. This severe phenotype could be partially rescued by the addition of a transgene carrying two copies of the normal human methylmalonyl-CoA mutase locus. The “humanized” mice were smaller than control litter mates and had high levels of methylmalonic acid in their blood and tissues. This new transgenic MMA stop codon model mimics (at both the phenotypic and genotypic levels) the key features of the human MMA disorder. It will allow the trialing of pharmacological and, cell and gene therapies for the treatment of MMA and other human metabolic disorders caused by stop codon mutations. PMID:23024777

  16. An evaluation of new and established methods to determine T-DNA copy number and homozygosity in transgenic plants.

    PubMed

    Głowacka, Katarzyna; Kromdijk, Johannes; Leonelli, Lauriebeth; Niyogi, Krishna K; Clemente, Tom E; Long, Stephen P

    2016-04-01

    Stable transformation of plants is a powerful tool for hypothesis testing. A rapid and reliable evaluation method of the transgenic allele for copy number and homozygosity is vital in analysing these transformations. Here the suitability of Southern blot analysis, thermal asymmetric interlaced (TAIL-)PCR, quantitative (q)PCR and digital droplet (dd)PCR to estimate T-DNA copy number, locus complexity and homozygosity were compared in transgenic tobacco. Southern blot analysis and ddPCR on three generations of transgenic offspring with contrasting zygosity and copy number were entirely consistent, whereas TAIL-PCR often underestimated copy number. qPCR deviated considerably from the Southern blot results and had lower precision and higher variability than ddPCR. Comparison of segregation analyses and ddPCR of T1 progeny from 26 T0 plants showed that at least 19% of the lines carried multiple T-DNA insertions per locus, which can lead to unstable transgene expression. Segregation analyses failed to detect these multiple copies, presumably because of their close linkage. This shows the importance of routine T-DNA copy number estimation. Based on our results, ddPCR is the most suitable method, because it is as reliable as Southern blot analysis yet much faster. A protocol for this application of ddPCR to large plant genomes is provided.

  17. Muscle-directed gene therapy for phenylketonuria (PKU): Development of transgenic mice with muscle-specific phenylalanine hydroxylase expression

    SciTech Connect

    Harding, C.O.; Messing, A.; Wolff, J.A.

    1994-09-01

    Phenylketonuria (PKU) is an attractive target for gene therapy because of shortcomings in current therapy including lifelong commitment to a difficult and expensive diet, persistent mild cognitive deficits in some children despite adequate dietary therapy, and maternal PKU syndrome. Phenylalanine hydroxylase (PAH) is normally expressed only in liver, but we propose to treat PKU by introducing the gene for PAH into muscle. In order to evaluate both the safety and efficacy of this approach, we have a developed a trangenic mouse which expresses PAH in both cardiac and skeletal muscle. The transgene includes promoter and enhancer sequences from the mouse muscle creatine kinase (MCK) gene fused to the mouse liver PAH cDNA. Mice which have inherited the transgene are healthy, active, and do not exhibit any signs of muscle weakness or wasting. Ectopic PAH expression in muscle is not detrimental to the health, neurologic function, or reproduction of the mice. Pah{sup enu2} hyperphenylalaninemic mice, a model of human PAH deficiency, bred to carry the transgene have substantial PAH expression in cardiac and skeletal muscle but none in liver. Muscle PAH expression alone does not complement the hyperphenylalaninemic phenotype of Pah{sup enu2} mice. However, administration of reduced tetrahydrobiopterin to transgenic Pah{sup enu2} mice is associated with a 25% mean decrease in serum phenylalanine levels. We predict that ectopic expression of PAH in muscle along with adequate muscle supplies of reduced biopterin cofactor will decrease hyperphenylalaninemia in PKU.

  18. An AANAT/ASMT transgenic animal model constructed with CRISPR/Cas9 system serving as the mammary gland bioreactor to produce melatonin-enrich milk in sheep.

    PubMed

    Ma, Teng; Tao, Jingli; Yang, Minghui; He, Changjiu; Tian, Xiuzhi; Zhang, Xiaosheng; Zhang, Jinlong; Deng, Shoulong; Feng, Jianzhong; Zhang, Zhenzhen; Wang, Jing; Ji, Pengyun; Song, Yukun; He, Pingli; Han, Hongbing; Fu, Juncai; Lian, Zhengxing; Liu, Guoshi

    2017-03-08

    Melatonin as a potent antioxidant exhibits important nutritional and medicinal values. To produce melatonin-enriching milk will benefit to the consumers. In this study, a sheep bioreactor which generates melatonin-enriching milk has been successfully developed by the technology that combined CRISPR/Cas9 system and microinjection. The AANAT and ASMT were cloned from pineal gland of Dorper sheep (Ovis aries). The in vitro studies found that AANAT and ASMT were successfully transferred to the mammary epithelial cell lines and significantly increased melatonin production in the culture medium compared to the non-transgenic cell lines. In addition, the Cas9 mRNA, sgRNA and the linearized vectors pBC1-AANAT and pBC1-ASMT were co-injected into the cytoplasm of pronuclear embryos which were implanted into ewes by oviducts transferring. Thirty four transgenic sheep were generated with the transgenic positive rate being roughly 35% which were identified by southern-blot and sequencing. Seven carried transgenic AANAT, two carried ASMT and 25 carried both of AANAT and ASMT genes. RT-PCR and western-blot demonstrated that the lambs expressed these genes in their mammary epithelial cells and these animals produced melatonin-enriched milk. This is the first report to show a functional AANAT and ASMT transgenic animal model which produce significantly high levels of melatonin milk compared to their wild type counterparts. The advanced technologies used in the study laid a foundation for generating large transgenic livestock, for example the cows, which can produce high level of melatonin milk. This article is protected by copyright. All rights reserved.

  19. [Effect of transgenic insect-resistant rice on biodiversity].

    PubMed

    Zhang, Lei; Zhu, Zhen

    2011-05-01

    Rice is the most important food crops in maintaining food security in China. The loss of China's annual rice production caused by pests is over ten million tons. Present studies showed that the transgenic insect-resistant rice can substantially reduce the application amount of chemical pesticides. In the case of no pesticide use, the pest density in transgenic rice field is significantly lower than that in non-transgenic field, and the neutral insects and natural enemies of pests increased significantly, indicating that the ecological environment and biodiversity toward the positive direction. The gene flow frequency from transgenic rice is dramatically reduced with the distance increases, reaching less than 0.01% at the distance of 6.2 m. Application of transgenic insect-resistant rice in China has an important significance for ensuring food security, maintaining sustainable agricultural development, and protecting the ecological environment and biodiversity. This review summarized the research progress in transgenic insect-resistant rice and its effect on biodiversity. The research directions and development trends of crop pest controlling in future are discussed. These help to promote better use of transgenic insect-resistant rice.

  20. Acute acetaminophen toxicity in transgenic mice with elevated hepatic glutathione.

    PubMed

    Rzucidlo, S J; Bounous, D I; Jones, D P; Brackett, B G

    2000-06-01

    Previous studies demonstrated that elevation of hepatic glutathione (GSH) concentrations protect against acetaminophen (APAP) hepatotoxicity in mice. Employing transgenic mice overexpressing glutathione synthetase, this study was conducted to determine if sustained elevation of hepatic GSH concentrations could ameliorate or prevent APAP toxicity. International Cancer Research transgenic mouse males and matched (ie same strain, sex, and age) control nontransgenic mice were pretreated ip with GSH synthetase substrate gamma-glutamylcysteinyl ethyl ester (gamma-GCE) or with saline. After a 16-h fast, mice received a single dose of 500 mg APAP/kg bw in saline ip and were sacrificed 4 h later. Other mice similarly pretreated were killed without APAP challenge. The elevated GSH concentrations in transgenic mice livers did not lessen APAP hepatotoxicity. Instead higher degrees of hepatotoxicity and nephrotoxicity were observed in transgenic mice than in controls as indicated by higher serum alanine aminotransferase activity and more severe histopathological lesions in transgenic mice livers and kidneys. Pretreatment with gamma-GCE did not affect either initial or post-APAP treatment tissue GSH concentrations or observed degrees of toxicity. Detection of a higher level of serum APAP in transgenic mice and the histopathological lesions found in transgenic mice kidneys together with no observable nephrotoxicity in control mice indicated early kidney damage in transgenic mice. Our findings suggest that high levels of GSH-APAP conjugates resulting from increased GSH concentrations in the livers of transgenic mice caused rapid kidney damage. Compromised excretory ability may have caused retention of APAP, which, in effect, elicited higher hepatotoxicity than that observed in nontransgenic mice.

  1. Transgenic fish systems and their application in ecotoxicology.

    PubMed

    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.

  2. Efficient CRISPR-mediated gene targeting and transgene replacement in the beetle Tribolium castaneum.

    PubMed

    Gilles, Anna F; Schinko, Johannes B; Averof, Michalis

    2015-08-15

    Gene-editing techniques are revolutionizing the way we conduct genetics in many organisms. The CRISPR/Cas nuclease has emerged as a highly versatile, efficient and affordable tool for targeting chosen sites in the genome. Beyond its applications in established model organisms, CRISPR technology provides a platform for genetic intervention in a wide range of species, limited only by our ability to deliver it to cells and to select mutations efficiently. Here, we test the CRISPR technology in an emerging insect model and pest, the beetle Tribolium castaneum. We use simple assays to test CRISPR/Cas activity, we demonstrate efficient expression of guide RNAs and Cas9 from Tribolium U6 and hsp68 promoters and we test the efficiency of knockout and knock-in approaches in Tribolium. We find that 55-80% of injected individuals carry mutations (indels) generated by non-homologous end joining, including mosaic bi-allelic knockouts; 71-100% carry such mutations in their germ line and transmit them to the next generation. We show that CRISPR-mediated gene knockout of the Tribolium E-cadherin gene causes defects in dorsal closure, which is consistent with RNAi-induced phenotypes. Homology-directed knock-in of marker transgenes was observed in 14% of injected individuals and transmitted to the next generation by 6% of injected individuals. Previous work in Tribolium mapped a large number of transgene insertions associated with developmental phenotypes and enhancer traps. We present an efficient method for re-purposing these insertions, via CRISPR-mediated replacement of these transgenes by new constructs.

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

    PubMed Central

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

    2016-01-01

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

  4. Characterization of Fluorescent Eye Markers for Mammalian Transgenic Studies

    PubMed Central

    Cornett, Jonathan C.; Landrette, Sean F.; Xu, Tian

    2011-01-01

    Genotyping mice by DNA based methods is both laborious and costly. As an alternative, we systematically examined fluorescent proteins expressed in the lens as transgenic markers for mice. A set of eye markers has been selected such that double and triple transgenic animals can be visually identified and that fluorescence intensity in the eyes can be used to distinguish heterozygous from homozygous mice. Taken together, these eye markers dramatically reduce the time and cost of genotyping transgenics and empower analysis of genetic interaction. PMID:22216292

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

    PubMed

    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.

  6. Diploid potato (Solanum tuberosum L.) as a model crop to study transgene expression.

    PubMed

    Nadolska-Orczyk, Anna; Pietrusinska, Aleksandra; Binka-Wyrwa, Agnieszka; Kuc, Dominik; Orczyk, Wacław

    2007-01-01

    This paper presents a method of Agrobacterium-mediated transformation for two diploid breeding lines of potato, and gives a detailed analysis of reporter gene expression. In our lab, these lines were also used to obtain tetraploid somatic hybrids. We tested four newly prepared constructs based on the pGreen vector system containing the selection gene nptII or bar under the 35S or nos promoter. All these vectors carried gus under 35S. We also tested the pDM805 vector, with the bar and gus genes respectively under the Ubi1 and Act1 promoters, which are strong for monocots. The selection efficiency (about 17%) was highest in the stem and leaf explants after transformation with pGreen where nptII was under 35S. About half of the selected plants were confirmed via PCR and Southern blot analysis to be transgenic and, depending on the combination, 0 to 100% showed GUS expression. GUS expression was strongest in multi-copy transgenic plants where gus was under Act1. The same potato lines carrying multi-copy bar under Ubi1 were also highly resistant to the herbicide Basta. The suggestion of using Agrobacterium-mediated transformation of diploid lines of potato as a model crop is discussed herein.

  7. Expression Cloning and Production of Human Heavy-Chain-Only Antibodies from Murine Transgenic Plasma Cells

    PubMed Central

    Drabek, Dubravka; Janssens, Rick; de Boer, Ernie; Rademaker, Rik; Kloess, Johannes; Skehel, John; Grosveld, Frank

    2016-01-01

    Several technologies have been developed to isolate human antibodies against different target antigens as a source of potential therapeutics, including hybridoma technology, phage and yeast display systems. For conventional antibodies, this involves either random pairing of VH and variable light (VL) domains in combinatorial display libraries or isolation of cognate pairs of VH and VL domains from human B cells or from transgenic mice carrying human immunoglobulin loci followed by single-cell sorting, single-cell RT-PCR, and bulk cloning of isolated natural VH–VL pairs. Heavy-chain-only antibodies (HCAbs) that naturally occur in camelids require only heavy immunoglobulin chain cloning. Here, we present an automatable novel, high-throughput technology for rapid direct cloning and production of fully human HCAbs from sorted population of transgenic mouse plasma cells carrying a human HCAb locus. Utility of the technique is demonstrated by isolation of diverse sets of sequence unique, soluble, high-affinity influenza A strain X-31 hemagglutinin-specific HCAbs. PMID:28066429

  8. FUNCTIONAL PROTEOME OF MACROPHAGE CARRIED NANOFORMULATED ANTIRETROVIRAL THERAPY DEMONSTRATES ENHANCED PARTICLE CARRYING CAPACITY

    PubMed Central

    Martinez-Skinner, Andrea L.; Veerubhotla, Ram S.; Liu, Han; Xiong, Huangui; Yu, Fang; McMillan, JoEllyn M.; Gendelman, Howard E.

    2013-01-01

    Our laboratory has pioneered the development of long-acting nanoformulations of antiretroviral therapy (nanoART). NanoART serves to improve drug compliance, toxicities, and access to viral reservoirs. These all function to improve treatment of human immunodeficiency virus (HIV) infection. Formulations are designed to harness the carrying capacities of mononuclear phagocytes (MP; monocytes and macrophages) and to use these cells as Trojan horses for drug delivery. Such a drug distribution system limits ART metabolism and excretion while facilitating access to viral reservoirs. Our prior works demonstrated a high degree of nanoART sequestration in macrophage recycling endosomes with broad and sustained drug tissue biodistribution and depots with limited untoward systemic toxicities. Despite such benefits, the effects of particle carriage on the cells’ functional capacities remained poorly understood. Thus, we employed pulsed stable isotope labeling of amino acids in cell culture to elucidate the macrophage proteome and assess any alterations in cellular functions that would affect cell-drug carriage and release kinetics. NanoART-MP interactions resulted in the induction of a broad range of activation-related proteins that can enhance phagocytosis, secretory functions, and cell migration. Notably, we now demonstrate that particle-cell interactions serve to enhance drug loading while facilitating drug tissue depots and transportation. PMID:23544708

  9. Down with DON: Strategies for precise transgene delivery and rnai-based suppression of fusarium

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Transgenic strategies can effectively supplement other methods for controlling Fusarium head blight (FHB). Impediments to deploying FHB-resistant transgenic barley include a long time-frame for creating and testing transgenes in barley, imprecise transgene insertions that lead to unstable gene expre...

  10. 32 CFR 552.103 - Requirements for carrying and use.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... Requirements for carrying and use. Persons legally authorized to possess firearms, ammunition, knives (with... engaged in hunting or shooting. Knives will be carried in a sheath or scabbard worn in a clearly visible manner. Commanders may authorize the carrying of a privately-owned knife with a blade over 3 inches...

  11. 32 CFR 552.103 - Requirements for carrying and use.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... Requirements for carrying and use. Persons legally authorized to possess firearms, ammunition, knives (with... engaged in hunting or shooting. Knives will be carried in a sheath or scabbard worn in a clearly visible manner. Commanders may authorize the carrying of a privately-owned knife with a blade over 3 inches...

  12. 32 CFR 552.103 - Requirements for carrying and use.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... Requirements for carrying and use. Persons legally authorized to possess firearms, ammunition, knives (with... engaged in hunting or shooting. Knives will be carried in a sheath or scabbard worn in a clearly visible manner. Commanders may authorize the carrying of a privately-owned knife with a blade over 3 inches...

  13. 21 CFR 880.6900 - Hand-carried stretcher.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Hand-carried stretcher. 880.6900 Section 880.6900 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED... Devices § 880.6900 Hand-carried stretcher. (a) Identification. A hand-carried stretcher is a...

  14. 21 CFR 880.6900 - Hand-carried stretcher.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Hand-carried stretcher. 880.6900 Section 880.6900 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED... Devices § 880.6900 Hand-carried stretcher. (a) Identification. A hand-carried stretcher is a...

  15. 21 CFR 880.6900 - Hand-carried stretcher.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Hand-carried stretcher. 880.6900 Section 880.6900 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED... Devices § 880.6900 Hand-carried stretcher. (a) Identification. A hand-carried stretcher is a...

  16. 21 CFR 880.6900 - Hand-carried stretcher.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Hand-carried stretcher. 880.6900 Section 880.6900 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED... Devices § 880.6900 Hand-carried stretcher. (a) Identification. A hand-carried stretcher is a...

  17. 21 CFR 880.6900 - Hand-carried stretcher.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Hand-carried stretcher. 880.6900 Section 880.6900 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED... Devices § 880.6900 Hand-carried stretcher. (a) Identification. A hand-carried stretcher is a...

  18. 46 CFR 111.105-35 - Vessels carrying coal.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 46 Shipping 4 2014-10-01 2014-10-01 false Vessels carrying coal. 111.105-35 Section 111.105-35...-GENERAL REQUIREMENTS Hazardous Locations § 111.105-35 Vessels carrying coal. (a) The following are Class II, Division 1, (Zone 10 or Z) locations on a vessel that carries coal: (1) The interior of each...

  19. 46 CFR 111.105-35 - Vessels carrying coal.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 46 Shipping 4 2013-10-01 2013-10-01 false Vessels carrying coal. 111.105-35 Section 111.105-35...-GENERAL REQUIREMENTS Hazardous Locations § 111.105-35 Vessels carrying coal. (a) The following are Class II, Division 1, (Zone 10 or Z) locations on a vessel that carries coal: (1) The interior of each...

  20. 46 CFR 111.105-35 - Vessels carrying coal.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... 46 Shipping 4 2012-10-01 2012-10-01 false Vessels carrying coal. 111.105-35 Section 111.105-35...-GENERAL REQUIREMENTS Hazardous Locations § 111.105-35 Vessels carrying coal. (a) The following are Class II, Division 1, (Zone 10 or Z) locations on a vessel that carries coal: (1) The interior of each...

  1. 46 CFR 111.105-35 - Vessels carrying coal.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 46 Shipping 4 2010-10-01 2010-10-01 false Vessels carrying coal. 111.105-35 Section 111.105-35...-GENERAL REQUIREMENTS Hazardous Locations § 111.105-35 Vessels carrying coal. (a) The following are Class II, Division 1, (Zone 10 or Z) locations on a vessel that carries coal: (1) The interior of each...

  2. 46 CFR 111.105-35 - Vessels carrying coal.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 46 Shipping 4 2011-10-01 2011-10-01 false Vessels carrying coal. 111.105-35 Section 111.105-35...-GENERAL REQUIREMENTS Hazardous Locations § 111.105-35 Vessels carrying coal. (a) The following are Class II, Division 1, (Zone 10 or Z) locations on a vessel that carries coal: (1) The interior of each...

  3. Polycythemia in transgenic mice expressing the human erythropoietin gene

    SciTech Connect

    Semenza, G.L.; Traystman, M.D.; Gearhart, J.D.; Antonarakis, S.E. )

    1989-04-01

    Erythropoietin is a glycoprotein hormone that regulates mammalian erythropoiesis. To study the expression of the human erythropoietin gene, EPO, 4 kilobases of DNA encompassing the gene with 0.4 kilobase of 5{prime} flanking sequence and 0.7 kilobase of 3{prime} flanking sequence was microinjected into fertilized mouse eggs. Transgenic mice were generated that are polycythemic, with increased erythrocytic indices in peripheral blood, increased numbers of erythroid precursors in hematopoietic tissue, and increased serum erythropoietin levels. Transgenic homozygotes show a greater degree of polycythemia than do heterozygotes as well as striking extramedullary erythropoiesis. Human erythropoietin RNA was found not only in fetal liver, adult liver, and kidney but also in all other transgenic tissues analyzed. Anemia induced increased human erythropoietin RNA levels in liver but not kidney. These transgenic mice represent a unique model of polycythemia due to increased erythropoietin levels.

  4. [Effects of agricultural activities and transgenic crops on agricultural biodiversity].

    PubMed

    Zhang, Xi-Tao; Luo, Hong-Bing; Li, Jun-Sheng; Huang, Hai; Liu, Yong-Bo

    2014-09-01

    Agricultural biodiversity is a key part of the ecosystem biodiversity, but it receives little concern. The monoculture, environmental pollution and habitat fragmentation caused by agricultural activities have threatened agricultural biodiversity over the past 50 years. To optimize agricultural management measures for crop production and environmental protection, we reviewed the effects of agricultural activities, including cultivation patterns, plastic mulching, chemical additions and the cultivation of transgenic crops, on agricultural biodiversity. The results showed that chemical pesticides and fertilizers had the most serious influence and the effects of transgenic crops varied with other factors like the specific transgene inserted in crops. The environmental risk of transgenic crops should be assessed widely through case-by-case methods, particularly its potential impacts on agricultural biodiversity. It is important to consider the protection of agricultural biodiversity before taking certain agricultural practices, which could improve agricultural production and simultaneously reduce the environmental impacts.

  5. Designer proton-channel transgenic algae for photobiological hydrogen production

    DOEpatents

    Lee, James Weifu [Knoxville, TN

    2011-04-26

    A designer proton-channel transgenic alga for photobiological hydrogen production that is specifically designed for production of molecular hydrogen (H.sub.2) through photosynthetic water splitting. The designer transgenic alga includes proton-conductive channels that are expressed to produce such uncoupler proteins in an amount sufficient to increase the algal H.sub.2 productivity. In one embodiment the designer proton-channel transgene is a nucleic acid construct (300) including a PCR forward primer (302), an externally inducible promoter (304), a transit targeting sequence (306), a designer proton-channel encoding sequence (308), a transcription and translation terminator (310), and a PCR reverse primer (312). In various embodiments, the designer proton-channel transgenic algae are used with a gas-separation system (500) and a gas-products-separation and utilization system (600) for photobiological H.sub.2 production.

  6. OPTIMAL BAND SELECTION OF HYPERSPECTRAL DATA FOR TRANSGENIC CORN IDENTIFICATION

    EPA Science Inventory

    Resistance development by insect pests to the insecticidal proteins expressed in transgenic crops would increase reliance on broad spectrum chemical insecticides subsequently reducing environmental quality and increasing worker exposure to toxic chemicals. An important component ...

  7. SCREENING OF TRANSGENIC ANTHURIUMS FOR BACTERIAL BLIGHT AND NEMATODE RESISTANCE

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Anthuriums exhibit limited resistance to bacterial blight caused by Xanthomonas axonopodis pv. dieffenbachiae and to the nematodes Radopholus simile and Meloidogyne javanica. Agrobacterium tumefaciens transformation of embryogenic calli with strains LBA4404, EHA105, and AGLO resulted in transgenic p...

  8. Strategies for metabolic pathway engineering with multiple transgenes.

    PubMed

    Bock, Ralph

    2013-09-01

    The engineering of metabolic pathways in plants often requires the concerted expression of more than one gene. While with traditional transgenic approaches, the expression of multiple transgenes has been challenging, recent progress has greatly expanded our repertoire of powerful techniques making this possible. New technological options include large-scale co-transformation of the nuclear genome, also referred to as combinatorial transformation, and transformation of the chloroplast genome with synthetic operon constructs. This review describes the state of the art in multigene genetic engineering of plants. It focuses on the methods currently available for the introduction of multiple transgenes into plants and the molecular mechanisms underlying successful transgene expression. Selected examples of metabolic pathway engineering are used to illustrate the attractions and limitations of each method and to highlight key factors that influence the experimenter's choice of the best strategy for multigene engineering.

  9. Neurodevelopmental effects of decabromodiphenyl ether (BDE-209) in APOE transgenic mice.

    PubMed

    Reverte, Ingrid; Domingo, José L; Colomina, Maria Teresa

    2014-01-01

    Exposure to low doses of neurotoxic pollutants during early brain development is a public health concern. Perinatal exposure to polybrominated diphenyl ethers (PBDEs) has been associated with neurodevelopmental effects in infants and long-term behavioral alterations in rodents. Decabromodiphenyl ether (BDE-209) is extensively used in the industry, with its potential risk to humans still under examination. In a previous study, we found that a single postnatal administration of BDE-209 impaired spatial learning in mice at 12 months of age, but a similar alteration was present in young mice carrying a specific genotype of apolipoprotein E (apoE). On the basis of our results, the main goal of the current investigation was to assess whether the same exposure to BDE-209 would affect the neurodevelopment of apoE transgenic mice. We used a functional observational battery (FOB) to evaluate the physical and neuromotor maturation of transgenic mice carrying different apoE polymorphisms (ε2, ε3, and ε4). On postnatal day 10, BDE-209 was administered orally at 0, 10 and 30 mg/kg and neurodevelopmental screening was carried out until postnatal day 36. We observed a subtle delay in eye opening in mice carrying the apoE4 genotype. Exposure to the high dose of BDE-209 retarded the eye opening of apoE2 mice, but no other developmental features were affected. The results indicate few effects of BDE-209 during development, while the vulnerability conferred by the apoE genotype may vary depending on age. Identifying relevant early gene-environment interactions is fundamental for a better understanding of adult health and disease.

  10. Expression of cyclin D1 in epithelial tissues of transgenic mice results in epidermal hyperproliferation and severe thymic hyperplasia.

    PubMed Central

    Robles, A I; Larcher, F; Whalin, R B; Murillas, R; Richie, E; Gimenez-Conti, I B; Jorcano, J L; Conti, C J

    1996-01-01

    To study the involvement of cyclin D1 in epithelial growth and differentiation and its putative role as an oncogene in skin, transgenic mice were developed carrying the human cyclin D1 gene driven by a bovine keratin 5 promoter. As expected, all squamous epithelia including skin, oral mucosa, trachea, vaginal epithelium, and the epithelial compartment of the thymus expressed aberrant levels of cyclin D1. The rate of epidermal proliferation increased dramatically in transgenic mice, which also showed basal cell hyperplasia. However, epidermal differentiation was unaffected, as shown by normal growth arrest of newborn primary keratinocytes in response to high extracellular calcium. Moreover, an unexpected phenotype was observed in the thymus. Transgenic mice developed a severe thymic hyperplasia that caused premature death due to cardio-respiratory failure within 4 months of age. By 14 weeks, the thymi of transgenic mice increased in weight up to 40-fold, representing 10% of total body weight. The hyperplastic thymi had normal histology revealing a well-differentiated cortex and medulla, which supported an apparently normal T-cell developmental program based on the distribution of thymocyte subsets. These results suggest that proliferation and differentiation of epithelial cells are under independent genetic controls in these organs and that cyclin D1 can modulate epithelial proliferation without altering the initiation of differentiation programs. No spontaneous development of epithelial tumors or thymic lymphomas was perceived in transgenic mice during their first 8 months of life, although they continue under observation. This model provides in vivo evidence of the action of cyclin D1 as a pure mediator of proliferation in epithelial cells. Images Fig. 1 Fig. 2 Fig. 3 PMID:8755527

  11. DNMT 1 maintains hypermethylation of CAG promoter specific region and prevents expression of exogenous gene in fat-1 transgenic sheep

    PubMed Central

    Yang, Chunrong; Shang, Xueying; Cheng, Lei; Yang, Lei; Liu, Xuefei; Bai, Chunling; Wei, Zhuying; Hua, Jinlian; Li, Guangpeng

    2017-01-01

    Methylation is an important issue in gene expression regulation and also in the fields of genetics and reproduction. In this study, we created fat-1 transgenic sheep, investigated the fine-mapping and the modulatory mechanisms of promoter methylation. Sheep fetal fibroblasts were transfected by pCAG-fat1-IRES-EGFP. Monoclonal cell line was screened as nuclear donor and carried out nuclear transfer (441 transgenic cloned embryos, 52 synchronism recipient sheep). Six offsprings were obtained. Expressions of exogenous genes fat-1 and EGFP were detectable in 10 examined tissues and upregulated omega-3 fatty acid content. Interestingly, more or less EGFP negative cells were detectable in the positive transgenic fetal skin cells. EGFP negative and positive cells were sorted by flow cytometry, and their methylation status in the whole promoter region (1701 nt) were investigated by bisulphate sequencing. The fine-mapping of methylation in CAG promoter were proposed. The results suggested that exogenous gene expression was determined by the methylation status from 721–1346 nt and modulated by methylation levels at 101, 108 and 115 nt sites in CAG promoter. To clarify the regulatory mechanism of methylation, examination of four DNA methyltransferases (DNMTs) demonstrated that hypermethylation of CAG promoter is mainly maintained by DNMT 1 in EGFP negative cells. Furthermore, investigation of the cell surface antigen CD34, CD45 and CD166 indicated that EGFP positive and negative cells belong to different types. The present study systematically clarified methylation status of CAG promoter in transgenic sheep and regulatory mechanism, which will provide research strategies for gene expression regulation in transgenic animals. PMID:28158319

  12. Transgenic expression of the N525S-tuberin variant in Tsc2 mutant (Eker) rats causes dominant embryonic lethality

    PubMed Central

    Shiono, Masatoshi; Kobayashi, Toshiyuki; Takahashi, Riichi; Ueda, Masatsugu; Ishioka, Chikashi; Hino, Okio

    2014-01-01

    The Tsc2 product, tuberin, negatively regulates the mTOR pathway. We have exploited the Eker (Tsc2-mutant) rat system to analyse various Tsc2 mutations. Here, we focus on the N525S-Tsc2 variant (NSM), which is known to cause distinct symptoms in patients even though normal suppression of mTOR is observed. Unexpectedly, we were repeatedly unable to generate viable rats carrying the NSM transgene. Genotypic analysis revealed that most of the embryos carrying the transgene died around embryonic day after 14.5—similar to the stage of lethality observed for Eker homozygotes. Thus, the NSM transgene appeared to have a dominant lethal effect in our rat model. Further, no significant differences were observed for various signal transduction molecules in transiently expressed NSM cells compared to WT. These results indicate that a non-mTOR pathway, critical for embryogenesis, is being regulated by tuberin, providing a link between tuberin expression and the severity of Tsc2 mutation-related pathogenesis. PMID:25088526

  13. Cell-type-specific activity of the human papillomavirus type 18 upstream regulatory region in transgenic mice and its modulation by tetradecanoyl phorbol acetate and glucocorticoids.

    PubMed Central

    Cid, A; Auewarakul, P; Garcia-Carranca, A; Ovseiovich, R; Gaissert, H; Gissmann, L

    1993-01-01

    The upstream regulatory region (URR) of human papillomavirus type 18 (HPV-18) harbors transcriptional promoter and enhancer elements which are thought to determine the cell-type specificity of the virus. In order to study the regulation of HPV-18 expression in vivo, we constructed transgenic mice carrying the bacterial lacZ gene under the control of the HPV-18 URR. Analysis of beta-galactosidase activity by histochemical staining of tissue sections of four independent transgenic mice showed that the viral promoter was specifically active in epithelial cells within a variety of organs (e.g., tongue, ovary, uterus, testis, and small intestine). Very strong staining was observed in newborn transgenic mice in contrast to a weak activity found during fetal life. Determination of beta-galactosidase activity in crude extracts from tissues of three lines of transgenic mice proved to be a useful tool for a quantitative analysis of transgene expression. In mice from two different transgenic lines treated with dexamethasone such measurements revealed a biphasic effect of the hormone on the activity of the enzyme in the stratified epithelium of the tongue (transient increase followed by a decrease). Northern (RNA) blot analysis showed similar changes in beta-galactosidase mRNA in that tissue. Treatment with tetradecanoyl phorbol acetate (TPA) led to a twofold increase in both enzymatic activity and mRNA levels. Finally, combined treatments with dexamethasone and TPA showed that both factors interfered with each other in their respective effects on transgene expression, suggesting that a cross-talk mechanism between transcription factors could be involved in the regulation of the HPV-18 URR. Images PMID:8411377

  14. Altered Epiphytic Colonization of Mannityl Opine-Producing Transgenic Tobacco Plants by a Mannityl Opine-Catabolizing Strain of Pseudomonas syringae

    PubMed Central

    Wilson, M.; Savka, M. A.; Hwang, I.; Farrand, S. K.; Lindow, S. E.

    1995-01-01

    The plasmid pYDH208, which confers the ability to catabolize the mannityl opines mannopine and agropine, was mobilized into the nonpathogenic Pseudomonas syringae strain Cit7. The growth of the mannityl opine-catabolizing strain Cit7(pYDH208) was compared with that of the near-isogenic non-opine-catabolizing strain Cit7xylE on leaves of wild-type tobacco (Nicotiana tabacum cv. Xanthi) and transgenic mannityl opine-producing tobacco plants (N. tabacum cv. Xanthi, line 2-26). The population size of Cit7(pYDH208) was significantly greater on the lower leaves of transgenic plants than on middle or upper leaves of those plants. The population size of Cit7(pYDH208) on lower leaves of transgenic plants was also significantly greater than the population size of Cit7xylE on similar leaves of wild-type plants. High-voltage paper electrophoresis demonstrated higher levels of mannityl opines in washings from lower- and mid-level leaves than in washings from upper-level leaves. The ability of Cit7(pYDH208) to catabolize mannityl opines in the carbon-limited phyllosphere increased the carrying capacity of the lower leaves of transgenic plants for Cit7(pYDH208). In coinoculations, the increase in the ratio of population sizes of Cit7(pYDH208) to Cit7xylE on transgenic plants was apparently due to a subtle difference in the growth rates of the two strains and to the difference in final population sizes. An ability to utilize additional carbon sources on the transgenic plants also enabled Cit7(pYDH208) to achieve a higher degree of coexistence with Cit7xylE on transgenic plants than on wild-type plants. This supports the hypothesis that the level of coexistence between epiphytic bacterial populations can be altered through nutritional resource partitioning. PMID:16535040

  15. Transgenic cloned sheep overexpressing ovine toll-like receptor 4.

    PubMed

    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.

  16. Lectin cDNA and transgenic plants derived therefrom

    DOEpatents

    Raikhel, Natasha V.

    1994-01-04

    Transgenic plants containing cDNA encoding Gramineae lectin are described. The plants preferably contain cDNA coding for barley lectin and store the lectin in the leaves. The transgenic plants, particularly the leaves exhibit insecticidal and fungicidal properties. GOVERNMENT RIGHTS This application was funded under Department of Energy Contract DE-AC02-76ER01338. The U.S. Government has certain rights under this application and any patent issuing thereon.

  17. Lectin cDNA and transgenic plants derived therefrom

    DOEpatents

    Raikhel, N.V.

    1994-01-04

    Transgenic plants containing cDNA encoding Gramineae lectin are described. The plants preferably contain cDNA coding for barley lectin and store the lectin in the leaves. The transgenic plants, particularly the leaves exhibit insecticidal and fungicidal properties. GOVERNMENT RIGHTS This application was funded under Department of Energy Contract DE-AC02-76ER01338. The U.S. Government has certain rights under this application and any patent issuing thereon. .

  18. [Research progress of transgenic Drosophila model of Alzheimer disease].

    PubMed

    Tan, Yan; Ji, Yu-Bin; Zhao, Jian

    2013-03-01

    Alzheimer disease (AD) is a common neurodegenerative disease. Drosophila has been regard as one of the ideal models for Alzheimer because of its unique advantage on genetic manipulation. AD transgenic drosophila models not only help to elucidate the pathogenesis of Alzheimer disease, but also provide potential screening models for drugs to treat the disease. In this review, we summarize the recent research progress using AD transgenic drosophila.

  19. Dihydrofolate Reductase and Thymidylate Synthase Transgenes Resistant to Methotrexate Interact to Permit Novel Transgene Regulation*

    PubMed Central

    Rushworth, David; Mathews, Amber; Alpert, Amir; Cooper, Laurence J. N.

    2015-01-01

    Methotrexate (MTX) is an anti-folate that inhibits de novo purine and thymidine nucleotide synthesis. MTX induces death in rapidly replicating cells and is used in the treatment of multiple cancers. MTX inhibits thymidine synthesis by targeting dihydrofolate reductase (DHFR) and thymidylate synthase (TYMS). The use of MTX to treat cancer also causes bone marrow suppression and inhibits the immune system. This has led to the development of an MTX-resistant DHFR, DHFR L22F, F31S (DHFRFS), to rescue healthy cells. 5-Fluorouracil-resistant TYMS T51S, G52S (TYMSSS) is resistant to MTX and improves MTX resistance of DHFRFS in primary T cells. Here we find that a known mechanism of MTX-induced increase in DHFR expression persists with DHFRFS and cis-expressed transgenes. We also find that TYMSSS expression of cis-expressed transgenes is similarly decreased in an MTX-inducible manner. MTX-inducible changes in DHFRFS and TYMSSS expression changes are lost when both genes are expressed together. In fact, expression of the DHFRFS and TYMSSS cis-expressed transgenes becomes correlated. These findings provide the basis for an unrecognized post-transcriptional mechanism that functionally links expression of DHFR and TYMS. These findings were made in genetically modified primary human T cells and have a clear potential for use in clinical applications where gene expression needs to be regulated by drug or maintained at a specific expression level. We demonstrate a potential application of this system in the controlled expression of systemically toxic cytokine IL-12. PMID:26242737

  20. [Obtaining transgenic rice plants and their progenies using Agrobacterium tumefaciens].

    PubMed

    Yin, Z C; Yang, F; Xu, Y; Li, B J

    1998-12-01

    Rice (Oriza sativa L.) suspension cells of Taipei 309 were co-cultivated with A. tumefaciens stran EHA101 harbouring binary vector pBYT2 for 3 days in the presence of vir inducer, 100 mumol/L acetosyringone (AS). After 2 months of continuous selection, 17 stable hygromycin-resistant, GUS-positive calli were recovered from 364 suspension cell clusters co-cultivated with A. tumefaciens. 10 putative transgenic R0 plants obtained from 8 tansformed calli and their progenies were analyzed for the integration and expression of foreign genes. Southern blot analysis of R0 and R1 generations indicated that foreign genes had been stably integrated in the genome of transgenic rice and sexually transmitted. One of the transgenic lines showed 5 copies of T-DNA integration, while the others had only one copy. Histochemical staining observation and fluorometric assay of GUS activity in transgenic rice cells and plants showed ubiquitin promoter from maize was highly effective in driving the expression of gus reporter gene in transgenic rice cells. GUS protein and its activity were also investigated through ndPAGE-X-Gluc staining assay, and it was found that the GUS protein in transgenic rice cells was smaller in size than the standard GUS protein (Sigma Co. G0786) but as large as that from E.coli HB101 (pBI121). This study suggested that Agrobacterium-mediated transformation of plant is an efficient and reliable method to introduce foreign genes into rice.

  1. Enhanced malignant tumorigenesis in Cdk4 transgenic mice.

    PubMed

    Miliani de Marval, Paula L; Macias, Everardo; Conti, Claudio J; Rodriguez-Puebla, Marcelo L

    2004-03-11

    In a previous study, we reported that overexpression of cyclin-dependent kinase-4 (CDK4) in mouse epidermis results in epidermal hyperplasia, hypertrophy and severe dermal fibrosis. In this study, we have investigated the susceptibility to skin tumor formation by forced expression of CDK4. Skin tumors from transgenic mice showed a dramatic increase in the rate of malignant progression to squamous cell carcinomas (SCC) in an initiation-promotion protocol. Histopathological analysis of papillomas from transgenic mice showed an elevated number of premalignant lesions characterized by dysplasia and marked atypia. Interestingly, transgenic mice also developed tumors in initiated but not promoted skin, demonstrating that CDK4 replaced the action of tumor promoters. These results suggest that expression of cyclin D1 upon ras activation synergizes with CDK4 overexpression. However, cyclin D1 transgenic mice and double transgenic mice for cyclin D1 and CDK4 did not show increased malignant progression in comparison to CDK4 transgenic mice. Biochemical analysis of tumors showed that CDK4 sequesters the CDK2 inhibitors p27Kip1 and p21Cip1, suggesting that indirect activation of CDK2 plays an important role in tumor development. These results indicate that, contrary to the general assumption, the catalytic subunit, CDK4, has higher oncogenic activity than cyclin D1, revealing a potential use of CDK4 as therapeutic target.

  2. Neurologic and motor dysfunctions in APP transgenic mice

    PubMed Central

    Lalonde, Robert; Fukuchi, Ken-ichiro; Strazielle, Catherine

    2012-01-01

    The discovery of gene mutations underlying autosomal dominant Alzheimer’s disease has enabled researchers to reproduce several hallmarks of this disorder in transgenic mice, notably the formation of Aβ plaques in brain and cognitive deficits. APP transgenic mutants have also been investigated with respect to survival rates, neurologic functions, and motor coordination, which are all susceptible to alteration in Alzheimer dementia. Several transgenic lines expressing human mutated or wild-type APP had higher mortality rates than non-transgenic controls with or without the presence of Aβ plaques. Mortality rates were also elevated in APP transgenic mice with vascular amyloid accumulation, thereby implicating cerebrovascular factors in the precocious death observed in all APP transgenic models. In addition, myoclonic jumping has been described in APP mutants, together with seizure activity, abnormal limb-flexion and paw-clasping reflexes, and motor coordination deficits. The neurologic signs resemble the myoclonic movements, epileptic seizures, pathological reflexes, and gait problems observed in late-stage Alzheimer’s disease. PMID:23089603

  3. Gene expression profiling of R6/2 transgenic mice with different CAG repeat lengths reveals genes associated with disease onset and progression in Huntington's disease.

    PubMed

    Tang, Bin; Seredenina, Tamara; Coppola, Giovanni; Kuhn, Alexandre; Geschwind, Daniel H; Luthi-Carter, Ruth; Thomas, Elizabeth A

    2011-06-01

    R6/2 transgenic mice with expanded CAG repeats (>300) have a surprisingly prolonged disease progression and longer lifespan than prototypical parent R6/2 mice (carrying 150 CAGs); however, the mechanism of this phenotype amelioration is unknown. We compared gene expression profiles in the striatum of R6/2 transgenic mice carrying ~300 CAG repeats (R6/2(Q300) transgenic mice) to those carrying ~150 CAG repeats (R6/2(Q150) transgenic mice) and littermate wildtype controls in order to identify genes that may play determinant roles in the time course of phenotypic expression in these mice. Of the top genes showing concordant expression changes in the striatum of both R6/2 lines, 85% were decreased in expression, while discordant expression changes were observed mostly for genes upregulated in R6/2(Q300) transgenic mice. Upregulated genes in the R6/2(Q300) mice were associated with the ubiquitin ligase complex, cell adhesion, protein folding, and establishment of protein localization. We qPCR-validated increases in expression of genes related to the latter category, including Lrsam1, Erp29, Nasp, Tap1, Rab9b, and Pfdn5 in R6/2(Q300) mice, changes that were not observed in R6/2 mice with shorter CAG repeats, even in late stages (i.e., 12 weeks of age). We further tested Lrsam1 and Erp29, the two genes showing the greatest upregulation in R6/2(Q300) transgenic mice, for potential neuroprotective effects in primary striatal cultures overexpressing a mutated human huntingtin (htt) fragment. Overexpression of Lrsam1 prevented the loss of NeuN-positive cell bodies in htt171-82Q cultures, concomitant with a reduction of nuclear htt aggregates. Erp29 showed no significant effects in this model. This is consistent with the distinct pattern of htt inclusion localization observed in R6/2(Q300) transgenic mice, in which smaller cytoplasmic inclusions represent the major form of insoluble htt in the cell, as opposed to large nuclear inclusions observed in R6/2(Q150) transgenic mice

  4. Development of real time PCR assays for detection and quantification of transgene DNA of a Bacillus thuringiensis (Bt) corn hybrid in soil samples.

    PubMed

    Zhu, Bin; Ma, Bao-Luo; Blackshaw, Robert E

    2010-10-01

    Real time PCR assays were developed to detect and quantify the transgene DNA of a commercially released Bacillus thuringiensis (Bt) corn (Zea mays L.) hybrid (DKC42-23), which was derived from the event MON863 and also carried a neomycin phosphotransferase gene (the nptII gene). We applied the real time PCR assays to investigate the persistence of the transgene DNA in a field trial grown with DKC42-23 over 3 years, in combination with bacterial natural transformation. The results showed that under continuous cultivation of DKC42-23, its transgene DNA was detectable in the field plots all year around. Meanwhile, when soil DNA extracts from DKC42-23 plots were used as donor in bacterial natural transformation, successful recovery of kanamycin resistant (Km(R)) transformants indicated that the nptII gene carried by DKC42-23 could be taken up and integrated into naturally competent Pseudomonas stutzeri pMR7 cells, leading to the restoration of the antibiotic resistance of P. stutzeri pMR7. However, after the cultivation of a soybean line in the same plots for the subsequent growing season, the presence of transgene DNA of DKC42-23 was reduced to undetectable levels at the end of that growing season. Therefore, existing corn-soybean crop rotation practices reduce the availability of transgene DNA in soil and thus minimize the risks that might be attributable to horizontal gene transfer. The real time PCR assays are useful for investigating the persistence of transgene DNA derived from the MON863 event in soil environments.

  5. Transgenic crops coping with water scarcity.

    PubMed

    Cominelli, Eleonora; Tonelli, Chiara

    2010-11-30

    Water scarcity is a serious problem that will be exacerbated by global climate change. Massive quantities of water are used in agriculture, and abiotic stresses, especially drought and increased salinity, are primary causes of crop loss worldwide. Various approaches may be adopted to consume less water in agriculture, one of them being the development of plants that use less water yet maintain high yields in conditions of water scarcity. In recent years several molecular networks concerned with stress perception, signal transduction and stress responses in plants have been elucidated. Consequently, engineering some of the genes involved in these mechanisms promises to enhance plant tolerance to stresses and in particular increase their water use efficiency. Here we review the various approaches used so far to produce transgenic plants having improved tolerance to abiotic stresses, and discuss criteria for choosing which genes to work on (functional and regulatory genes) and which gene expression promoters (constitutive, inducible, and cell-specific) have been used to obtain successful results.

  6. Epigenetic Regulation of Intronic Transgenes in Arabidopsis

    PubMed Central

    Osabe, Kenji; Harukawa, Yoshiko; Miura, Saori; Saze, Hidetoshi

    2017-01-01

    Defense mechanisms of plant genomes can epigenetically inactivate repetitive sequences and exogenous transgenes. Loss of mutant phenotypes in intronic T-DNA insertion lines by interaction with another T-DNA locus, termed T-DNA suppression, has been observed in Arabidopsis thaliana, although the molecular basis of establishment and maintenance of T-DNA suppression is poorly understood. Here we show that maintenance of T-DNA suppression requires heterochromatinisation of T-DNA sequences and the nuclear proteins, INCREASED IN BONSAI METHYLATION 2 (IBM2) and ENHANCED DOWNY MILDEW 2 (EDM2), which prevent ectopic 3′ end processing of mRNA in atypically long introns containing T-DNA sequences. Initiation of T-DNA suppression is mediated by the canonical RdDM pathway after hybridisation of two T-DNA strains, accompanied by DNA hypermethylation of T-DNA sequences in the F1 generation. Our results reveal the presence of a genome surveillance mechanism through genome hybridisation that masks repetitive DNAs intruding into transcription units. PMID:28338020

  7. Evaluating cerebellar functions using optogenetic transgenic mice

    NASA Astrophysics Data System (ADS)

    Welsh, John P.; Turecek, Josef; Turner, Eric E.

    2013-03-01

    We employed a transgenic mouse having conditional expression of ChR2(H134R) in neurons of the inferior olive to facilitate understanding of the role of electrical coupling and oscillation in central nervous system function. Two-photon excitation of ChR2-expressing neurons using 64 laser beams restricted to single inferior olive cell bodies depolarized neurons and evoked voltage deflections in neighboring neurons demonstrating electrical coupling. Broader illumination of neuronal ensembles using blue light induced an optical clamp of endogenous electrical rhythms in the inferior olive of acutely-prepared brain slices, which when applied in vivo directly modulated the local field potential activity and induced tremor. The experiments demonstrate novel methods to optically manipulate electrically coupled potentials and rhythmogenesis within a neuronal ensemble. From a functional perspective, the experiments shed light on the cellular and circuitry mechanisms of essential tremor, a prevalent neurological condition, by indicating time- and frequencydependence of tremor upon varying rhythms of inferior olive stimulation. The experiments indicate analog control of a brain rhythm that may be used to enhance our understanding of the functional consequences of central rhythmogenesis.

  8. AMPK: Lessons from transgenic and knockout animals

    PubMed Central

    Viollet, Benoit; Athea, Yoni; Mounier, Remi; Guigas, Bruno; Zarrinpashneh, Elham; Horman, Sandrine; Lantier, Louise; Hebrard, Sophie; Devin-Leclerc, Jocelyne; Beauloye, Christophe; Foretz, Marc; Andreelli, Fabrizio; Ventura-Clapier, Renee; Bertrand, Luc

    2009-01-01

    AMP-activated protein kinase (AMPK), a phylogenetically conserved serine/threonine protein kinase, has been proposed to function as a ‘fuel gauge’ to monitor cellular energy status in response to nutritional environmental variations. AMPK system is a regulator of energy balance that, once activated by low energy status, switches on ATP-producing catabolic pathways (such as fatty acid oxidation and glycolysis), and switches off ATP-consuming anabolic pathways (such as lipogenesis), both by short-term effect on phosphorylation of regulatory proteins and by long-term effect on gene expression. Numerous observations obtained with pharmacological activators and agents that deplete intracellular ATP have been supportive of AMPK playing a role in the control of energy metabolism but none of these studies have provided conclusive evidence. Relatively recent developments in our understanding of precisely how AMPK complexes might operate to control energy metabolism is due in part to the development of transgenic and knockout mouse models. Although there are inevitable caveats with genetic models, some important findings have emerged. In the present review, we discuss recent findings obtained from animal models with inhibition or activation of AMPK signaling pathway. PMID:19273052

  9. Stable transgene expression in primitive human CD34+ hematopoietic stem/progenitor cells, using the Sleeping Beauty transposon system.

    PubMed

    Sumiyoshi, Teiko; Holt, Nathalia G; Hollis, Roger P; Ge, Shundi; Cannon, Paula M; Crooks, Gay M; Kohn, Donald B

    2009-12-01

    Sleeping Beauty (SB) transposon-mediated integration has been shown to achieve long-term transgene expression in a wide range of host cells. In this study, we improved the SB transposon-mediated gene transfer system for transduction of human CD34(+) stem/progenitor cells by two approaches: (1) to increase the transposition efficacy, a hyperactive mutant of SB, HSB, was used; (2) to improve the expression of the SB transposase and the transgene cassette carried by the transposon, different viral and cellular promoters were evaluated. SB components were delivered in trans into the target cells by Nucleoporation. The SB transposon-mediated integration efficacy was assessed by integrated transgene (enhanced green fluorescent protein [eGFP]) expression both in vitro and in vivo. In purified human cord blood CD34(+) cells, HSB achieved long-term transgene expression in nearly 7-fold more cells than the original SB transposase. Significantly brighter levels of eGFP expression (5-fold) were achieved with the human elongation factor 1alpha (EF1-alpha) promoter in Jurkat human T cells, compared with that achieved with the modified myeloproliferative sarcoma virus long terminal repeat enhancer-promoter (MNDU3); in contrast, the MNDU3 promoter expressed eGFP at the highest level in K-562 myeloid cells. In human CD34(+) cord blood cells studied under conditions directing myeloid differentiation, the highest transgene integration and expression were achieved using the EF1-alpha promoter to express the SB transposase combined with the MNDU3 promoter to express the eGFP reporter. Stable transgene expression was achieved at levels up to 27% for more than 4 weeks of culture after improved gene transfer to CD34(+) cells (average, 17%; n = 4). In vivo studies evaluating engraftment and differentiation of the SB-modified human CD34(+) cells demonstrated that SB-modified human CD34(+) cells engrafted in NOD/SCID/gamma chain(null) (NSG) mice and differentiated into multilineage cell

  10. Complete sequence of the IncT-type plasmid pT-OXA-181 carrying the blaOXA-181 carbapenemase gene from Citrobacter freundii.

    PubMed

    Villa, Laura; Carattoli, Alessandra; Nordmann, Patrice; Carta, Claudio; Poirel, Laurent

    2013-04-01

    The gene encoding the carbapenemase OXA-181 (an OXA-48 variant) was identified from a Citrobacter freundii isolate coproducing NDM-1. The whole sequence of plasmid pT-OXA-181 bearing the blaOXA-181 gene was determined and revealed a 84-kb mobilizable but non-self-conjugative IncT-type plasmid. It totally differs from the 7.6-kb ColE-type and blaOXA-181-bearing plasmid recently identified in a Klebsiella pneumoniae isolate. However, in both plasmids, insertion sequence ISEcp1 might have played a role in acquisition of the blaOXA-181 gene.

  11. Agrobacterium tumefaciens-mediated creeping bentgrass (Agrostis stolonifera L.) transformation using phosphinothricin selection results in a high frequency of single-copy transgene integration.

    PubMed

    Luo, H; Hu, Q; Nelson, K; Longo, C; Kausch, A P; Chandlee, J M; Wipff, J K; Fricker, C R

    2004-04-01

    Genetic transformation of creeping bentgrass mediated by Agrobacterium tumefaciens has been achieved. Embryogenic callus initiated from seeds (cv. Penn-A-4) was infected with an A. tumefaciens strain (LBA4404) harboring a super-binary vector that contained an herbicide-resistant bar gene driven either by the CaMV 35S promoter or a rice ubiquitin promoter. Plants were regenerated from 219 independent transformation events. The overall stable transformation efficiency ranged from 18% to 45%. Southern blot and genetic analysis confirmed transgene integration in the creeping bentgrass genome and normal transmission and stable expression of the transgene in the T1 generation. All independent transformation events carried one to three copies of the transgene, and a majority (60-65%) contained only a single copy of the foreign gene with no apparent rearrangements. We report here the successful use of Agrobacterium for the large-scale production of transgenic creeping bentgrass plants with a high frequency of a single-copy transgene insertion that exhibit stable inheritance patterns.

  12. Comparative study of transgenic and non-transgenic maize (Zea mays) flours commercialized in Brazil, focussing on proteomic analyses.

    PubMed

    Vidal, Nádia; Barbosa, Herbert; Jacob, Silvana; Arruda, Marco

    2015-08-01

    Genetically modified foods are a major concern around the world due to the lack of information concerning their safety and health effects. This work evaluates differences, at the proteomic level, between two types of crop samples: transgenic (MON810 event with the Cry1Ab gene, which confers resistance to insects) and non-transgenic maize flour commercialized in Brazil. The 2-D DIGE technique revealed 99 differentially expressed spots, which were collected in 2-D PAGE gels and identified via mass spectrometry (nESI-QTOF MS/MS). The abundance of protein differences between the transgenic and non-transgenic samples could arise from genetic modification or as a result of an environmental influence pertaining to the commercial sample. The major functional category of proteins identified was related to disease/defense and, although differences were observed between samples, no toxins or allergenic proteins were found.

  13. Construction of Sterile ime1Δ-Transgenic Saccharomyces cerevisiae Wine Yeasts Unable To Disseminate in Nature▿

    PubMed Central

    Ramírez, Manuel; Ambrona, Jesús

    2008-01-01

    The use of new transgenic yeasts in industry carries a potential environmental risk because their dispersal, introducing new artificial genetic combinations into nature, could have unpredictable consequences. This risk could be avoided by using sterile transgenic yeasts that are unable to sporulate and mate with wild yeasts. These sterile yeasts would not survive the annual cyclic harvesting periods, being condemned to disappear in the wineries and vineyards in less than a year. We have constructed new ime1Δ wine yeasts that are unable to sporulate and mate, bear easy-to-detect genetic markers, and quickly disappear in grape must fermentation immediately after sporulation of the yeast population. These sterile yeasts maintained the same biotechnological properties as their parent yeasts without any detectable deleterious effect of the ime1Δ mutation. These yeasts are therefore interesting biotechnologically for food industry applications and for genetically modified microorganism environmental monitoring studies. PMID:18245242

  14. Efficient production by sperm-mediated gene transfer of human decay accelerating factor (hDAF) transgenic pigs for xenotransplantation

    PubMed Central

    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

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

    PubMed

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

    PubMed

    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.

  17. Phenotypic rescue by a bovine transgene in a Cu/Zn superoxide dismutase-null mutant of Drosophila melanogaster

    SciTech Connect

    Reveillaud, I.; Kongpachith, A.; Fleming, J.E.

    1994-02-01

    Null mutants for Cu/Zn superoxide dismutase (CuZnSOD) in Drosophila melanogaster are male sterile, have a greatly reduced adult life span, and are hypersensitive to paraquat. We have introduced a synthetic bovine CuZnSOD transgene under the transcriptional control of the D. melanogaster 5C actin promoter into a CuZnSOD-null mutant of D. melanogaster. This was carried out by P-element-mediated transformation of the Drosophila-bovine CuZnSOD transgene into a CuZnSOD{sup +} recipient strain followed by genetic crossing of the transgene into a strain carrying the CuZnSOD-null mutation, cSOD{sup n108}. The resulting transformants express bovine CuZnSOD exclusively to about 30% of normal Drosophila CuZnSOD levels. Expression of the Drosophila-bovine CuZnSOD transgene in the CuZnSOD-null mutant rescues male fertility and resistance to paraquat to apparently normal levels. However, adult life span is restored to only 30% of normal, and resistance to hyperoxia is 90% of that found in control flies. This striking differential restoration of pleiotropic phenotypes could be the result of a threshold of CuZnSOD expression necessary for normal male fertility and resistance to the toxicity of paraquat or hyperoxia which is lower than the threshold required to sustain a normal adult life span. Alternatively, the differential rescue of fertility, resistance to active oxygen, and life span might indicate different cell-specific transcriptional requirements for these functions which are normally provided by the control elements of the native CuZnSOD gene but are only partly compensated for by the transcriptional control elements of the actin 5C promoter. 29 refs., 5 figs., 1 tab.

  18. Monitoring of Venus transgenic cell migration during pregnancy in non-transgenic rabbits.

    PubMed

    Lipták, N; Hoffmann, O I; Kerekes, A; Iski, G; Ernszt, D; Kvell, K; Hiripi, L; Bősze, Z

    2017-04-01

    Cell transfer between mother and fetus were demonstrated previously in several species which possess haemochorial placenta (e.g. in humans, mice, rats, etc.). Here we report the assessment of fetal and maternal microchimerism in non-transgenic (non-TG) New Zealand white rabbits which were pregnant with transgenic (TG) fetuses and in non-TG newborns of TG does. The TG construct, including the Venus fluorophore cDNA driven by a ubiquitous cytomegalovirus enhancer, chicken ß-actin promoter (CAGGS), was previously integrated into the rabbit genome by Sleeping Beauty transposon system. Three different methods [fluorescence microscopy, flow cytometry and quantitative polymerase chain reaction (QPCR)] were employed to search for TG cells and gene products in blood and other tissues of non-TG rabbits. Venus positive peripheral blood mononuclear cells (PBMCs) were not detected in the blood of non-TG littermates or non-TG does by flow cytometry. Tissue samples (liver, kidney, skeletal and heart muscle) also proved to be Venus negative examined with fluorescence microscopy, while histology sections and PBMCs of TG rabbits showed robust Venus protein expression. In case of genomic DNA (gDNA) sourced from tissue samples of non-TG rabbits, CAGGS promoter-specific fragments could not be amplified by QPCR. Our data showed the lack of detectable cell transfer between TG and non-TG rabbits during gestation.

  19. [Analysis of transgenic and non-transgenic rice leaves using visible/near-infrared spectroscopy].

    PubMed

    Zhu, Wen-chao; Cheng, Fang

    2012-02-01

    Visible/near-infrared (Vis/NIR) spectroscopy was investigated for the fast discrimination of rice leaves with different genes and the determination of chlorophyll content. Least squares-support vector machines (LS-SVM) was employed to discriminate transgenic rice leaves from non-transgenic ones. The classification accuracy of calibration samples reached to 100%. Successive projections algorithm (SPA) was proposed to select effective wavelengths. SPA-LS-SVM discrimination model was performed, and the result indicated that an 87.27% recognition ratio was achieved using only 0.3% of total variables. The optimal performance of each quantification model was achieved after orthogonal signal correction (OSA). Performances treated by SPA were better than that of full-spectrum PLS, which indicated that SPA is a powerful way for effective wavelength selection. The best performance of quantification was obtained by SPA-LS-SVM model; with correlation coefficient (R) and root mean square error of prediction (RMSEP) being 0.902 2 and 1.312 1, respectively. Excellent classification and prediction precision were achieved. The overall results indicated that the new proposed SPA-LS-SVM is a powerful method for varieties recognition and SPAD prediction. This study supplied a new and alternative approach to the further application of Vis/NIR spectroscopy in on-field classification and monitoring.

  20. Survival of skin graft between transgenic cloned dogs and non-transgenic cloned dogs.

    PubMed

    Kim, Geon A; Oh, Hyun Ju; Kim, Min Jung; Jo, Young Kwang; Choi, Jin; Park, Jung Eun; Park, Eun Jung; Lim, Sang Hyun; Yoon, Byung Il; Kang, Sung Keun; Jang, Goo; Lee, Byeong Chun

    2014-01-01

    Whereas it has been assumed that genetically modified tissues or cells derived from somatic cell nuclear transfer (SCNT) should be accepted by a host of the same species, their immune compatibility has not been extensively explored. To identify acceptance of SCNT-derived cells or tissues, skin grafts were performed between cloned dogs that were identical except for their mitochondrial DNA (mtDNA) haplotypes and foreign gene. We showed here that differences in mtDNA haplotypes and genetic modification did not elicit immune responses in these dogs: 1) skin tissues from genetically-modified cloned dogs were successfully transplanted into genetically-modified cloned dogs with different mtDNA haplotype under three successive grafts over 63 days; and 2) non-transgenic cloned tissues were accepted into transgenic cloned syngeneic recipients with different mtDNA haplotypes and vice versa under two successive grafts over 63 days. In addition, expression of the inserted gene was maintained, being functional without eliciting graft rejection. In conclusion, these results show that transplanting genetically-modified tissues into normal, syngeneic or genetically-modified recipient dogs with different mtDNA haplotypes do not elicit skin graft rejection or affect expression of the inserted gene. Therefore, therapeutically valuable tissue derived from SCNT with genetic modification might be used safely in clinical applications for patients with diseased tissues.

  1. Behavioral Indicators of Drug Carrying in Open Spaces

    DTIC Science & Technology

    2015-04-30

    Naval Research Laboratory Washington, DC 20375-5320 NRL/MR/5508--15-9596 Behavioral Indicators of Drug Carrying in Open Spaces April 30, 2015...NUMBER OF PAGES 17. LIMITATION OF ABSTRACT Behavioral Indicators of Drug Carrying in Open Spaces Nathan C. Meehan and Michael McClary1 Naval Research ...identification of people carrying illegal drugs . Further research assessing the reliability and validity of the behaviors identified in this report are

  2. High performance pipelined multiplier with fast carry-save adder

    NASA Technical Reports Server (NTRS)

    Wu, Angus

    1990-01-01

    A high-performance pipelined multiplier is described. Its high performance results from the fast carry-save adder basic cell which has a simple structure and is suitable for the Gate Forest semi-custom environment. The carry-save adder computes the sum and carry within two gate delay. Results show that the proposed adder can operate at 200 MHz for a 2-micron CMOS process; better performance is expected in a Gate Forest realization.

  3. Hepatic Microenvironment Affects Oval Cell Localization in Albumin-Urokinase-Type Plasminogen Activator Transgenic Mice

    PubMed Central

    Braun, Kristin M.; Thompson, Anne W.; Sandgren, Eric P.

    2003-01-01

    Mice carrying an albumin-urokinase type plasminogen activator transgene (AL-uPA) develop liver disease secondary to uPA expression in hepatocytes. Transgene-expressing parenchyma is replaced gradually by clones of cells that have deleted transgene DNA and therefore are not subject to uPA-mediated damage. Diseased liver displays several abnormalities, including hepatocyte vacuolation and changes in nonparenchymal tissue. The latter includes increases in laminin protein within parenchyma and the appearance of cytokeratin 19-positive bile ductule-like cells (oval cells) both in portal regions and extending into the hepatic parenchyma. In this study, we subjected AL-uPA mice to two-thirds partial hepatectomy to identify the response of these livers to additional growth stimulation. We observed several changes in hepatic morphology. First, the oval cells increased in number and often formed ductules in the parenchyma. Second, this cellular change was accompanied by a further increase in laminin associated with single or clusters of oval cells. Third, desmin-positive Ito cells increased in number and maintained close association with oval cells. Fourth, these changes were localized precisely to uPA-expressing areas of liver. Regenerating clones of uPA-deficient cells appeared to be unaffected both by stromal and cellular alterations. Thus, additional growth stimulation of diseased uPA-expressing liver induces an oval cell-like response, as observed in other models of severe hepatic injury, but the localization of this response seems to be highly regulated by the hepatic microenvironment. PMID:12507902

  4. Genotoxicity of 3-methylcholanthrene in liver of transgenic big Blue mice.

    PubMed

    Rihn, B H; Bottin, M C; Coulais, C; Rouget, R; Monhoven, N; Baranowski, W; Edorh, A; Keith, G

    2000-01-01

    Transgenic mice provide a unique tool for studying the tissue specificity and mutagenic potential of chemicals. Because 3-methylcholanthrene (3MC) was found mutagenic in bacteria, clastogenic in bone marrow, and induces DNA adducts in animals, we were interested to determinine whether this xenobiotic provokes (1) cell proliferation, (2) transcriptional activity changes, (3) DNA adducts, and (4) hepatic mutations in transgenic Big Blue mice carrying the lambdaLIZ phage shuttle vector. Big Blue C57/Bl male mice were treated with a single intraperitoneal dose of 80 mg/kg 3MC for 1, 3, 6, 14, or 30 days. Cell proliferation was checked by 5-bromo-2-deoxyuridine labeling and immunohistochemical detection. The maximal increase of the mitotic index was evidenced after 3 days (2.9 times the control value; P < 0.01). The relative nucleus area, reflecting the transcriptional activity, was also the highest in the treated group after 3 days: 1.86 times the control value, on average (P < 0.01). Four major DNA adducts, determined according to the [(32)P]-postlabeling method, were evidenced in liver DNA of treated mice, 6 days after the treatment: the spot intensities increased in a time-dependent manner. The mutant frequency of liver DNA was the highest after 14 days: 20.3 +/- 2.9 x 10(-5) in the treated vs. 7.6 +/- 2.7 x 10(-5) in the control mice (P < 0.01). Sequencing of the lambda lacI mutant plaques showed mainly G:C --> T:A and C:G --> A:T transversions. In conclusion, 3MC at first induced nuclear enlargement and a slight increase of cell proliferation in liver, followed by parallel formation of DNA adducts and mutations. This study shows how transgenic models allow in vivo evaluation of mechanistically simultaneous endpoints.

  5. Expression of a Pim-1 transgene accelerates lymphoproliferation and inhibits apoptosis in lpr/lpr mice.

    PubMed Central

    Möröy, T; Grzeschiczek, A; Petzold, S; Hartmann, K U

    1993-01-01

    Transgenic mice expressing the Pim-1 kinase are predisposed to develop T-cell lymphomas with a long latency period of about 7-9 months. However, the exact functional basis of the oncogenic activity of Pim-1 remains obscure. C57BL/6 mice homozygous for the lpr mutation develop a well-described lymphoproliferative syndrome at about 26-30 weeks of age. This syndrome is characterized mainly by the accumulation of abnormal T cells in lymph nodes because of the lack of Fas receptor-induced apoptosis. We find that backcross of E mu-Pim-1 transgenics (mice with a transgene that carries the mouse Pim-1 gene under the transcriptional control of the immunoglobulin heavy chain gene enhancer E mu) into lpr/lpr mice results in strong acceleration of lymphoproliferation and dramatic enlargement of lymph nodes. In addition, we show here that cultured lymph node cells from E mu-Pim-1 lpr/lpr mice are rescued from rapid apoptosis that normally occurs in nontransgenic lpr cells in vitro. We also present evidence that CD4+/CD8+ double-positive thymocytes from lpr/lpr mice are sensitive to dexamethasone-induced apoptosis, although lpr/lpr mice lack the Fas receptor. In contrast, E mu-Pim-1 lpr/lpr animals show considerable protection from dexamethasone-induced apoptosis. These results show that Pim-1 can strongly accelerate lymphoproliferation through inhibition of apoptosis and thereby provide first insight into the functional basis for the oncogenic activity of Pim-1. Images Fig. 1 Fig. 3 PMID:7504280

  6. Agrobacterium-mediated transformation of polyploid cereals. The efficiency of selection and transgene expression in wheat.

    PubMed

    Przetakiewicz, Anna; Karaś, Agnieszka; Orczyk, Wacław; Nadolska-Orczyk, Anna

    2004-01-01

    Three combinations of Agrobacterium tumefaciens strains and vectors were used in the transformation of selected Polish wheat cultivars. The combinations were: two hypervirulent strains, AGL1, containing the pDM805 binary plasmid, and EHA101, containing pGAH; and the common Agro strain LBA4404, harboring the super-binary pTOK233 vector. pDM805 contained bar under the control of Ubi1 promoter, pGAH had nptII under nos, and pTOK233 had hpt under 35S. Additionally, pDM805 and pTOK233 carried the gus reporter gene under the Act1 promoter or 35S promoter, respectively. The highest selection rate was 12.6% and was obtained with EHA101(pGAH) on a kanamycin-containing medium. Sixty-five of the plants grown on that medium were PCR positive. The second best combination was LBA4404(pTOK233) and kanamycin selection, which gave an average transformation rate of 2.3%. Phosphinothricin selection gave 1.0% transformation efficiency, while hygromycin, depending on the strain/vector used, gave from 0.2 to 0.4%. PCR tests in T1 revealed that 67% of the lines showed a 3:1 segregation ratio, and 11% a 15:1 ratio, while in 22%, segregation was non-Mendelian. The high number of T0 transgenic plants containing one copy of the transgene was confirmed via Southern blot analysis. Kanamycin resistance in the T1 generation was very low; in some lines, all the progeny were kanamycin sensitive. GUS expression, only tested in young T1 plants, was in agreement with Mendelian segregation in three out of the twelve tested. The factors influencing the efficiency of selection and transgene expression are discussed in this paper.

  7. Expression of hepatitis B surface antigen in transgenic banana plants.

    PubMed

    Kumar, G B Sunil; Ganapathi, T R; Revathi, C J; Srinivas, L; Bapat, V A

    2005-10-01

    Embryogenic cells of bananan cv. Rasthali (AAB) have been transformed with the 's' gene of hepatitis B surface antigen (HBsAg) using Agrobacterium mediated transformation. Four different expression cassettes (pHBS, pHER, pEFEHBS and pEFEHER) were utilized to optimize the expression of HBsAg in banana. The transgenic nature of the plants and expression of the antigen was confirmed by PCR, Southern hybridization and reverse transcription (RT)-PCR. The expression levels of the antigen in the plants grown under in vitro conditions as well as the green house hardened plants were estimated by ELISA for all the four constructs. Maximum expression level of 38 ng/g F.W. of leaves was noted in plants transformed with pEFEHBS grown under in vitro conditions, whereas pHER transformed plants grown in the green house showed the maximum expression level of 19.92 ng/g F.W. of leaves. Higher monoclonal antibody binding of 67.87% of the antigen was observed when it was expressed with a C-terminal ER retention signal. The buoyant density in CsCl of HBsAg derived from transgenic banana leaves was determined and found to be 1.146 g/ml. HBsAg obtained from transgenic banana plants is similar to human serum derived one in buoyant density properties. The transgenic plants were grown up to maturity in the green house and the expression of HBsAg in the fruits was confirmed by RT-PCR. These transgenic plants were multiplied under in vitro using floral apex cultures. Attempts were also made to enhance the expression of HBsAg in the leaves of transgenic banana plants by wounding and/or treatment with plant growth regulators. This is the first report on the expression of HBsAg in transgenic banana fruits.

  8. Striatal neurochemical changes in transgenic models of Huntington's disease.

    PubMed

    Ariano, Marjorie A; Aronin, Neil; Difiglia, Marian; Tagle, Danilo A; Sibley, David R; Leavitt, Blair R; Hayden, Michael R; Levine, Michael S

    2002-06-15

    Transgenic mouse models of Huntington's disease (HD) were examined following the onset of overt behavioral symptoms. The HD transgenic mice demonstrated profound striatal losses in D1, D2, and D3 dopamine (DA) receptor proteins in comparison with their nonsymptomatic, age-matched littermate controls. In parallel, a robust increase in the striatal D5 DA receptor subtype occurred in the transgenic compared with the wild-type control mice. This receptor elevation was accompanied by heightened cyclic AMP levels, which may be induced by the adenylyl cyclase-linked D5 receptor. This is a unique result; normal striatal D5 protein levels are modest and not thought to contribute substantially to cyclic AMP-mediated DA signaling mechanisms. Simple compensatory up-regulation of D5 DA receptors in response to D1 receptor subtype loss does not explain our findings, because genetic inactivation of the D1 DA receptor does not alter levels of D5 DA receptor expression. Immunofluorescent detection of tyrosine hydroxylase showed that nigrostriatal DA containing terminals were reduced, further supporting that disturbances in DA signaling occurred in HD transgenic models. The substance P-containing striatal efferent pathway was more resistant to the HD mutation than met-enkephalin-producing striatal projection neurons in the transgenics, based on neuropeptide immunofluorescent staining. Analogous findings in multiple transgenic models suggest that these changes are due to the presence of the transgene and are not dependent on its composition, promotor elements, or mouse strain background. These findings suggest modifications in the striatal DA system and that its downstream signaling through cyclic AMP mechanisms is disrupted severely in HD following onset of motor symptoms.

  9. Gene–environment interactions influence ecological consequences of transgenic animals

    PubMed Central

    Sundström, L. F.; Lõhmus, M.; Tymchuk, W. E.; Devlin, Robert H.

    2007-01-01

    Production of transgenic animals has raised concern regarding their potential ecological impact should they escape or be released to the natural environment. This concern has arisen mainly from research on laboratory-reared animals and theoretical modeling exercises. In this study, we used biocontained naturalized stream environments and conventional hatchery environments to show that differences in phenotype between transgenic and wild genotypes depend on rearing conditions and, critically, that such genotype-by-environment interactions may influence subsequent ecological effects in nature. Genetically wild and growth hormone transgenic coho salmon (Oncorhynchus kisutch) were reared from the fry stage under either standard hatchery conditions or under naturalized stream conditions. When reared under standard hatchery conditions, the transgenic fish grew almost three times longer than wild conspecifics and had (under simulated natural conditions) stronger predation effects on prey than wild genotypes (even after compensation for size differences). In contrast, when fish were reared under naturalized stream conditions, transgenic fish were only 20% longer than the wild fish, and the magnitude of difference in relative predation effects was much reduced. These data show that genotype-by-environment interactions can influence the relative phenotype of transgenic and wild-type organisms and that extrapolations of ecological consequences from phenotypes developed in the unnatural laboratory environment may lead to an overestimation or underestimation of ecological risk. Thus, for transgenic organisms that may not be released to nature, the establishment of a range of highly naturalized environments will be critical for acquiring reliable experimental data to be used in risk assessments. PMID:17360448

  10. [Development of a hepatitis B virus carrier transgenic mice model].

    PubMed

    Caner, Müge; Arat, Sezen; Bircan, Rifat

    2008-01-01

    The studies for the development of transgenic mice models which provide important profits for the studies concerning immunopathogenesis of hepatitis B virus (HBV) infections are in progress since 20 years. For this purpose different lineages bearing whole HBV genome or selected viral genes have been developed and their usage in clarifying the HBV replication and pathogenesis mechanisms have been emphasized. The aim of this study was to develop and breed a HBV carrier mice model. In the study the full HBV genome has been transferred to mouse embryos by microinjection procedure. Following transgenic manipulation, the HBV carriers among the daughter mice have been detected by molecular methods in which HBV-DNA replication and expression have been shown. The manipulations for transgene transfers have been performed in TUBITAK Marmara Research Center Transgene Laboratory, Gebze, Istanbul. The HBV-DNA carrier mice have been demonstrated by polymerase chain reaction (PCR) using the DNA samples obtained from tail tissues and also by dot-blot hybridization of the mice sera. Integrated HBV-DNA has been detected by applying in-situ hybridization to the liver tissue sections. HBV-DNA expression has been shown by reverse transcriptase PCR method with total RNA molecules that have been isolated from the liver tissues of the HBV-DNA carrier mice. HBsAg has been detected in the liver by immunohistochemical method, and HBsAg and HBeAg have additionally been demonstrated by ELISA. HBV genome, expression of the genome and the expression products have been determined in approximately 10% of the mice of which HBV-DNA have been transferred. By inbreeding heterozygote carrier mice, homozygote HBV transgenic mice line have been obtained. These HBV transgenic mice are the first lineages developed in our country. It is hopefully thought that this HBV carrier transgenic mouse model may contribute to the studies on the pathogenesis of HBV infections which are important health problems in the

  11. Challenges in predicting the evolutionary maintenance of a phage transgene

    PubMed Central

    2014-01-01

    Background In prior work, a phage engineered with a biofilm-degrading enzyme (dispersin B) cleared artificial, short-term biofilms more fully than the phage lacking the enzyme. An unresolved question is whether the transgene will be lost or maintained during phage growth – its loss would limit the utility of the engineering. Broadly supported evolutionary theory suggests that transgenes will be lost through a ‘tragedy of the commons’ mechanism unless the ecology of growth in biofilms meets specific requirements. We test that theory here. Results Functional properties of the transgenic phage were identified. Consistent with the previous study, the dispersin phage was superior to unmodified phage at clearing short term biofilms grown in broth, shown here to be an effect attributable to free enzyme. However, the dispersin phage was only marginally better than control phages on short term biofilms in minimal media and was no better than control phages in clearing long term biofilms. There was little empirical support for the tragedy of the commons framework despite a strong theoretical foundation for its supposed relevance. The framework requires that the transgene imposes an intrinsic cost, yet the transgene was intrinsically neutral or beneficial when expressed from one part of the phage genome. Expressed from a different part of the genome, the transgene did behave as if intrinsically costly, but its maintenance did not benefit from spatially structured growth per se – violating the tragedy framework. Conclusions Overall, the transgene was beneficial under many conditions, but no insight to its maintenance was attributable to the established evolutionary framework. The failure likely resides in system details that would be used to parameterize the models. Our study cautions against naive applications of evolutionary theory to synthetic biology, even qualitatively. PMID:25126112

  12. 25 CFR 23.51 - Grant carry-over authority.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... restricted by appropriation, and contingent upon satisfactory program evaluations from the appropriate area or agency office for an existing program, grantees are authorized to carry over unliquidated...

  13. 43 CFR 20.511 - Carrying of firearms.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 43 Public Lands: Interior 1 2011-10-01 2011-10-01 false Carrying of firearms. 20.511 Section 20... CONDUCT Other Employee Conduct Provisions § 20.511 Carrying of firearms. Employees, except those specifically designated to perform enforcement, police or other official duties requiring the use of...

  14. 43 CFR 20.511 - Carrying of firearms.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 43 Public Lands: Interior 1 2014-10-01 2014-10-01 false Carrying of firearms. 20.511 Section 20... CONDUCT Other Employee Conduct Provisions § 20.511 Carrying of firearms. Employees, except those specifically designated to perform enforcement, police or other official duties requiring the use of...

  15. 43 CFR 20.511 - Carrying of firearms.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 43 Public Lands: Interior 1 2013-10-01 2013-10-01 false Carrying of firearms. 20.511 Section 20... CONDUCT Other Employee Conduct Provisions § 20.511 Carrying of firearms. Employees, except those specifically designated to perform enforcement, police or other official duties requiring the use of...

  16. 43 CFR 20.511 - Carrying of firearms.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... 43 Public Lands: Interior 1 2012-10-01 2011-10-01 true Carrying of firearms. 20.511 Section 20.511... Other Employee Conduct Provisions § 20.511 Carrying of firearms. Employees, except those specifically designated to perform enforcement, police or other official duties requiring the use of firearms,...

  17. 43 CFR 20.511 - Carrying of firearms.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 43 Public Lands: Interior 1 2010-10-01 2010-10-01 false Carrying of firearms. 20.511 Section 20... CONDUCT Other Employee Conduct Provisions § 20.511 Carrying of firearms. Employees, except those specifically designated to perform enforcement, police or other official duties requiring the use of...

  18. 49 CFR 177.870 - Regulations for passenger carrying vehicles.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... PUBLIC HIGHWAY Regulations Applying to Hazardous Material on Motor Vehicles Carrying Passengers for Hire... apply to vehicles transporting passengers and property. (b) No Class 1 (explosive) materials or other hazardous materials on passenger-carrying vehicles, exceptions. No hazardous materials except...

  19. 47 CFR 76.1617 - Initial must-carry notice.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 47 Telecommunication 4 2014-10-01 2014-10-01 false Initial must-carry notice. 76.1617 Section 76.1617 Telecommunication FEDERAL COMMUNICATIONS COMMISSION (CONTINUED) BROADCAST RADIO SERVICES MULTICHANNEL VIDEO AND CABLE TELEVISION SERVICE Notices § 76.1617 Initial must-carry notice. (a) Within 60...

  20. 47 CFR 76.1617 - Initial must-carry notice.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 47 Telecommunication 4 2011-10-01 2011-10-01 false Initial must-carry notice. 76.1617 Section 76.1617 Telecommunication FEDERAL COMMUNICATIONS COMMISSION (CONTINUED) BROADCAST RADIO SERVICES MULTICHANNEL VIDEO AND CABLE TELEVISION SERVICE Notices § 76.1617 Initial must-carry notice. (a) Within 60...

  1. 47 CFR 76.1617 - Initial must-carry notice.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 47 Telecommunication 4 2013-10-01 2013-10-01 false Initial must-carry notice. 76.1617 Section 76.1617 Telecommunication FEDERAL COMMUNICATIONS COMMISSION (CONTINUED) BROADCAST RADIO SERVICES MULTICHANNEL VIDEO AND CABLE TELEVISION SERVICE Notices § 76.1617 Initial must-carry notice. (a) Within 60...

  2. 47 CFR 76.1617 - Initial must-carry notice.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... 47 Telecommunication 4 2012-10-01 2012-10-01 false Initial must-carry notice. 76.1617 Section 76.1617 Telecommunication FEDERAL COMMUNICATIONS COMMISSION (CONTINUED) BROADCAST RADIO SERVICES MULTICHANNEL VIDEO AND CABLE TELEVISION SERVICE Notices § 76.1617 Initial must-carry notice. (a) Within 60...

  3. 49 CFR 177.870 - Regulations for passenger carrying vehicles.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... PUBLIC HIGHWAY Regulations Applying to Hazardous Material on Motor Vehicles Carrying Passengers for Hire § 177.870 Regulations for passenger carrying vehicles. (a) Vehicles transporting passengers and property... apply to vehicles transporting passengers and property. (b) No Class 1 (explosive) materials or...

  4. 25 CFR 23.51 - Grant carry-over authority.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 25 Indians 1 2014-04-01 2014-04-01 false Grant carry-over authority. 23.51 Section 23.51 Indians... Uniform Grant Administration Provisions and Requirements § 23.51 Grant carry-over authority. Unless restricted by appropriation, and contingent upon satisfactory program evaluations from the appropriate...

  5. 25 CFR 11.444 - Carrying concealed weapons.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 25 Indians 1 2014-04-01 2014-04-01 false Carrying concealed weapons. 11.444 Section 11.444 Indians... ORDER CODE Criminal Offenses § 11.444 Carrying concealed weapons. A person who goes about in public places armed with a dangerous weapon concealed upon his or her person is guilty of a misdemeanor...

  6. 25 CFR 11.444 - Carrying concealed weapons.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 25 Indians 1 2010-04-01 2010-04-01 false Carrying concealed weapons. 11.444 Section 11.444 Indians... ORDER CODE Criminal Offenses § 11.444 Carrying concealed weapons. A person who goes about in public places armed with a dangerous weapon concealed upon his or her person is guilty of a misdemeanor...

  7. 25 CFR 11.444 - Carrying concealed weapons.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 25 Indians 1 2012-04-01 2011-04-01 true Carrying concealed weapons. 11.444 Section 11.444 Indians... ORDER CODE Criminal Offenses § 11.444 Carrying concealed weapons. A person who goes about in public places armed with a dangerous weapon concealed upon his or her person is guilty of a misdemeanor...

  8. 25 CFR 11.444 - Carrying concealed weapons.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 25 Indians 1 2013-04-01 2013-04-01 false Carrying concealed weapons. 11.444 Section 11.444 Indians... ORDER CODE Criminal Offenses § 11.444 Carrying concealed weapons. A person who goes about in public places armed with a dangerous weapon concealed upon his or her person is guilty of a misdemeanor...

  9. 25 CFR 11.444 - Carrying concealed weapons.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 25 Indians 1 2011-04-01 2011-04-01 false Carrying concealed weapons. 11.444 Section 11.444 Indians... ORDER CODE Criminal Offenses § 11.444 Carrying concealed weapons. A person who goes about in public places armed with a dangerous weapon concealed upon his or her person is guilty of a misdemeanor...

  10. Training to Increase Safe Tray Carrying among Cocktail Servers

    ERIC Educational Resources Information Center

    Scherrer, Megan D.; Wilder, David A.

    2008-01-01

    We evaluated the effects of training on proper carrying techniques among 3 cocktail servers to increase safe tray carrying on the job and reduce participants' risk of developing musculoskeletal disorders. As participants delivered drinks to their tables, their finger, arm, and neck positions were observed and recorded. Each participant received…

  11. 25 CFR 23.51 - Grant carry-over authority.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 25 Indians 1 2011-04-01 2011-04-01 false Grant carry-over authority. 23.51 Section 23.51 Indians BUREAU OF INDIAN AFFAIRS, DEPARTMENT OF THE INTERIOR HUMAN SERVICES INDIAN CHILD WELFARE ACT General and Uniform Grant Administration Provisions and Requirements § 23.51 Grant carry-over authority....

  12. 25 CFR 23.51 - Grant carry-over authority.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 25 Indians 1 2013-04-01 2013-04-01 false Grant carry-over authority. 23.51 Section 23.51 Indians BUREAU OF INDIAN AFFAIRS, DEPARTMENT OF THE INTERIOR HUMAN SERVICES INDIAN CHILD WELFARE ACT General and Uniform Grant Administration Provisions and Requirements § 23.51 Grant carry-over authority....

  13. 25 CFR 23.51 - Grant carry-over authority.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 25 Indians 1 2012-04-01 2011-04-01 true Grant carry-over authority. 23.51 Section 23.51 Indians BUREAU OF INDIAN AFFAIRS, DEPARTMENT OF THE INTERIOR HUMAN SERVICES INDIAN CHILD WELFARE ACT General and Uniform Grant Administration Provisions and Requirements § 23.51 Grant carry-over authority....

  14. 19 CFR 122.88 - Aircraft carrying domestic (stopover) passengers.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 19 Customs Duties 1 2011-04-01 2011-04-01 false Aircraft carrying domestic (stopover) passengers... Passengers § 122.88 Aircraft carrying domestic (stopover) passengers. Airlines that commingle domestic... continuing on another aircraft to a second U.S. destination) with international passengers who are...

  15. 19 CFR 122.88 - Aircraft carrying domestic (stopover) passengers.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 19 Customs Duties 1 2014-04-01 2014-04-01 false Aircraft carrying domestic (stopover) passengers... Passengers § 122.88 Aircraft carrying domestic (stopover) passengers. Airlines that commingle domestic... continuing on another aircraft to a second U.S. destination) with international passengers who are...

  16. 19 CFR 122.88 - Aircraft carrying domestic (stopover) passengers.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 19 Customs Duties 1 2010-04-01 2010-04-01 false Aircraft carrying domestic (stopover) passengers... Passengers § 122.88 Aircraft carrying domestic (stopover) passengers. Airlines that commingle domestic... continuing on another aircraft to a second U.S. destination) with international passengers who are...

  17. 19 CFR 122.88 - Aircraft carrying domestic (stopover) passengers.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 19 Customs Duties 1 2012-04-01 2012-04-01 false Aircraft carrying domestic (stopover) passengers... Passengers § 122.88 Aircraft carrying domestic (stopover) passengers. Airlines that commingle domestic... continuing on another aircraft to a second U.S. destination) with international passengers who are...

  18. 46 CFR 122.340 - Vessels carrying vehicles.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... 46 Shipping 4 2012-10-01 2012-10-01 false Vessels carrying vehicles. 122.340 Section 122.340... Miscellaneous Operating Requirements § 122.340 Vessels carrying vehicles. (a) Automobiles or other vehicles must... vehicles freely in the event of fire or other disaster. The decks, where necessary, must be...

  19. 46 CFR 185.340 - Vessels carrying vehicles.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 46 Shipping 7 2013-10-01 2013-10-01 false Vessels carrying vehicles. 185.340 Section 185.340... TONS) OPERATIONS Miscellaneous Operating Requirements § 185.340 Vessels carrying vehicles. (a) Automobiles or other vehicles must be stowed in such a manner as to permit both passengers and crew to get...

  20. 46 CFR 122.340 - Vessels carrying vehicles.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 46 Shipping 4 2010-10-01 2010-10-01 false Vessels carrying vehicles. 122.340 Section 122.340... Miscellaneous Operating Requirements § 122.340 Vessels carrying vehicles. (a) Automobiles or other vehicles must... vehicles freely in the event of fire or other disaster. The decks, where necessary, must be...

  1. 46 CFR 185.340 - Vessels carrying vehicles.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 46 Shipping 7 2014-10-01 2014-10-01 false Vessels carrying vehicles. 185.340 Section 185.340... TONS) OPERATIONS Miscellaneous Operating Requirements § 185.340 Vessels carrying vehicles. (a) Automobiles or other vehicles must be stowed in such a manner as to permit both passengers and crew to get...

  2. 46 CFR 185.340 - Vessels carrying vehicles.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 46 Shipping 7 2010-10-01 2010-10-01 false Vessels carrying vehicles. 185.340 Section 185.340... TONS) OPERATIONS Miscellaneous Operating Requirements § 185.340 Vessels carrying vehicles. (a) Automobiles or other vehicles must be stowed in such a manner as to permit both passengers and crew to get...

  3. 46 CFR 185.340 - Vessels carrying vehicles.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 46 Shipping 7 2011-10-01 2011-10-01 false Vessels carrying vehicles. 185.340 Section 185.340... TONS) OPERATIONS Miscellaneous Operating Requirements § 185.340 Vessels carrying vehicles. (a) Automobiles or other vehicles must be stowed in such a manner as to permit both passengers and crew to get...

  4. 46 CFR 122.340 - Vessels carrying vehicles.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 46 Shipping 4 2014-10-01 2014-10-01 false Vessels carrying vehicles. 122.340 Section 122.340... Miscellaneous Operating Requirements § 122.340 Vessels carrying vehicles. (a) Automobiles or other vehicles must... vehicles freely in the event of fire or other disaster. The decks, where necessary, must be...

  5. 46 CFR 185.340 - Vessels carrying vehicles.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... 46 Shipping 7 2012-10-01 2012-10-01 false Vessels carrying vehicles. 185.340 Section 185.340... TONS) OPERATIONS Miscellaneous Operating Requirements § 185.340 Vessels carrying vehicles. (a) Automobiles or other vehicles must be stowed in such a manner as to permit both passengers and crew to get...

  6. Neutralizing antibodies against rotavirus produced in transgenically labelled purple tomatoes.

    PubMed

    Juárez, Paloma; Presa, Silvia; Espí, Joaquín; Pineda, Benito; Antón, María T; Moreno, Vicente; Buesa, Javier; Granell, Antonio; Orzaez, Diego

    2012-04-01

    Edible fruits are inexpensive biofactories for human health-promoting molecules that can be ingested as crude extracts or partially purified formulations. We show here the production of a model human antibody for passive protection against the enteric pathogen rotavirus in transgenically labelled tomato fruits. Transgenic tomato plants expressing a recombinant human immunoglobulin A (hIgA_2A1) selected against the VP8* peptide of rotavirus SA11 strain were obtained. The amount of hIgA_2A1 protein reached 3.6 ± 0.8% of the total soluble protein in the fruit of the transformed plants. Minimally processed fruit-derived products suitable for oral intake showed anti-VP8* binding activity and strongly inhibited virus infection in an in vitro virus neutralization assay. In order to make tomatoes expressing hIgA_2A1 easily distinguishable from wild-type tomatoes, lines expressing hIgA_2A1 transgenes were sexually crossed with a transgenic tomato line expressing the genes encoding Antirrhinum majus Rosea1 and Delila transcription factors, which confer purple colour to the fruit. Consequently, transgenically labelled purple tomato fruits expressing hIgA_2A1 have been developed. The resulting purple-coloured extracts from these fruits contain high levels of recombinant anti-rotavirus neutralizing human IgA in combination with increased amounts of health-promoting anthocyanins.

  7. Transgenic Expression of Dentin Phosphoprotein Inhibits Skeletal Development

    PubMed Central

    Zhang, H.; Liu, P.; Wang, S.; Liu, C.; Jani, P.; Lu, Y.; Qin, C.

    2016-01-01

    Dentin sialophosphoprotein (DSPP) is proteolytically processed into an NH2-terminal fragment called dentin sialoprotein (DSP) and a COOH-terminal fragment known as dentin phosphoprotein (DPP). These two fragments are believed to perform distinct roles in formation of bone and dentin. To investigate the functions of DPP in skeletal development, we generated transgenic mice to overexpress hemagglutinin (HA)-tagged DPP under the control of a 3.6 kb type I collagen (Col1a1) promoter (designated as Col1a1-HA-DPP). The Col1a1-HA-DPP transgenic mice were significantly smaller by weight, had smaller skeletons and shorter long bones than their wild type littermates, as demonstrated by X-ray radiography. They displayed reduced trabecular bone formation and narrower zones of proliferative and hypertrophic chondrocytes in the growth plates of the long bones. Histological analyses showed that the transgenic mice had reduced cell proliferation in the proliferating zone, but lacked obvious defects in the chondrocyte differentiation. In addition, the transgenic mice with a high level of transgene expression developed spontaneous long bone fractures. In conclusion, overexpressing DPP inhibited skeletal development, suggesting that the balanced actions between the NH2- and COOH-terminal fragments of DSPP may be required for normal skeletal development. PMID:26972716

  8. Generation of transgenic marmosets expressing genetically encoded calcium indicators

    PubMed Central

    Park, Jung Eun; Zhang, Xian Feng; Choi, Sang-Ho; Okahara, Junko; Sasaki, Erika; Silva, Afonso C.

    2016-01-01

    Chronic monitoring of neuronal activity in the living brain with optical imaging techniques became feasible owing to the continued development of genetically encoded calcium indicators (GECIs). Here we report for the first time the successful generation of transgenic marmosets (Callithrix jacchus), an important nonhuman primate model in neurophysiological research, which were engineered to express the green fluorescent protein (GFP)-based family of GECIs, GCaMP, under control of either the CMV or the hSyn promoter. High titer lentiviral vectors were produced, and injected into embryos collected from donor females. The infected embryos were then transferred to recipient females. Eight transgenic animals were born and shown to have stable and functional GCaMP expression in several different tissues. Germline transmission of the transgene was confirmed in embryos generated from two of the founder transgenic marmosets that reached sexual maturity. These embryos were implanted into six recipient females, three of which became pregnant and are in advanced stages of gestation. We believe these transgenic marmosets will be invaluable non-human primate models in neuroscience, allowing chronic in vivo monitoring of neural activity with functional confocal and multi-photon optical microscopy imaging of intracellular calcium dynamics. PMID:27725685

  9. Developing transgenic Anopheles mosquitoes for the sterile insect technique.

    PubMed

    Nolan, Tony; Papathanos, Philippos; Windbichler, Nikolai; Magnusson, Kalle; Benton, Jason; Catteruccia, Flaminia; Crisanti, Andrea

    2011-01-01

    In the last 10 years the availability of the genome sequence of Anopheles gambiae and the development of a transgenic technology for several species of Anopheles mosquitoes have, in combination, helped in enabling us to gain several insights into the biology of these mosquitoes that is relevant to their capacity as vectors of the malaria parasite. While this information is anticipated to inform many novel vector control strategies, the technique most likely to benefit in the near future from the availability of a reliable transgenic technology is the sterile insect technique (SIT), which relies on releasing large numbers of sterile insects to compete for mates in the wild, leading to population suppression. Although SIT has been proven to work reliably for many insects, the construction of suitable strains, and induction of sterility, has until now been a laborious process, combining classical genetics with radiation-induced sterility. Using transgenesis to create strains of Anopheles suitable for SIT could potentially offer several advantages over current approaches, in that the basic design of transgenic constructs designed for other insects should be rapidly transferable to mosquitoes, and induction of sterility as a product of the transgenic modification could obviate the requirement for radiation and its associated deleterious effects. In this paper the progress of different transgenic approaches in constructing tools for SIT will be reviewed.

  10. Identification and quantification of anthocyanins in transgenic purple tomato.

    PubMed

    Su, Xiaoyu; Xu, Jianteng; Rhodes, Davina; Shen, Yanting; Song, Weixing; Katz, Benjamin; Tomich, John; Wang, Weiqun

    2016-07-01

    Anthocyanins are natural pigments derived from the phenylpropanoid pathway. Most tomatoes produce little anthocyanins, but the transgenic purple tomato biosynthesizes a high level of anthocyanins due to expression of two transcription factors (Del and Ros1). This study was to identify and quantify anthocyanins in this transgenic tomato line. Seven anthocyanins, including two new anthocyanins [malvidin-3-(p-coumaroyl)-rutinoside-5-glucoside and malvidin-3-(feruloyl)-rutinoside-5-glucoside], were identified by LC-MS/MS. Petunidin-3-(trans-coumaroyl)-rutinoside-5-glucoside and delphinidin-3-(trans-coumaroyl)-rutinoside-5-glucoside were the most abundant anthocyanins, making up 86% of the total anthocyanins. Compared to undetectable anthocyanins in the wild type, the contents of anthocyanins in the whole fruit, peel, and flesh of the Del/Ros1-transgenic tomato were 5.2±0.5, 5.1±0.5, and 5.8±0.3g/kg dry matter, respectively. Anthocyanins were undetectable in the seeds of both wide-type and transgenic tomato lines. Such novel and high levels of anthocyanins obtained in this transgenic tomato may provide unique functional products with potential health benefits.

  11. CCK Response Deficiency in Synphilin-1 Transgenic Mice.

    PubMed

    Smith, Wanli W; Smith, Megan; Yang, Dejun; Choi, Pique P; Moghadam, Alexander; Li, Tianxia; Moran, Timothy H

    2015-01-01

    Previously, we have identified a novel role for the cytoplasmic protein, synphilin-1(SP1), in the controls of food intake and body weight in both mice and Drosophila. Ubiquitous overexpression of human SP1 in brain neurons in transgenic mice results in hyperphagia expressed as an increase in meal size. However, the mechanisms underlying this action of SP1 remain to be determined. Here we investigate a potential role for altered gut feedback signaling in the effects of SP1 on food intake. We examined responses to peripheral administration of cholecytokinin (CCK), amylin, and the glucagon like peptide-1 (GLP-1) receptor agonist, exendin-4. Intraperitoneal administration of CCK at doses ranging from 1-10 nmol/kg significantly reduced glucose intake in wild type (WT) mice, but failed to affect intake in SP1 transgenic mice. Moreover, there was a significant attenuation of CCK-induced c-Fos expression in the dorsal vagal complex in SP1 transgenic mice. In contrast, WT and SP1 transgenic mice were similarly responsive to both amylin and exendin-4 treatment. These studies demonstrate that SP1 results in a CCK response deficiency that may contribute to the increased meal size and overall hyperphagia in synphillin-1 transgenic mice.

  12. Bioaccessibility of carotenoids from transgenic provitamin A biofortified sorghum.

    PubMed

    Lipkie, Tristan E; De Moura, Fabiana F; Zhao, Zuo-Yu; Albertsen, Marc C; Che, Ping; Glassman, Kimberly; Ferruzzi, Mario G

    2013-06-19

    Biofortified sorghum (Sorghum bicolor (L.) Moench) lines are being developed to target vitamin A deficiency in Sub-Saharan Africa, but the delivery of provitamin A carotenoids from such diverse germplasms has not been evaluated. The purpose of this study was to screen vectors and independent transgenic events for the bioaccessibility of provitamin A carotenoids using an in vitro digestion model. The germplasm background and transgenic sorghum contained 1.0-1.5 and 3.3-14.0 μg/g β-carotene equivalents on a dry weight basis (DW), respectively. Test porridges made from milled transgenic sorghum contained up to 250 μg of β-carotene equivalents per 100 g of porridge on a fresh weight basis (FW). Micellarization efficiency of all-trans-β-carotene was lower (p < 0.05) from transgenic sorghum (1-5%) than from null/nontransgenic sorghum (6-11%) but not different between vector constructs. Carotenoid bioaccessibility was significantly improved (p < 0.05) by increasing the amount of coformulated lipid in test porridges from 5% w/w to 10% w/w. Transgenic sorghum event Homo188-A contained the greatest bioaccessible β-carotene content, with a 4-8-fold increase from null/nontransgenic sorghum. While the bioavailability and bioconversion of provitamin A carotenoids from these grains must be confirmed in vivo, these data support the notion that biofortification of sorghum can enhance total and bioaccessible provitamin A carotenoid levels.

  13. Generation of Transgenic Monkeys with Human Inherited Genetic Disease

    PubMed Central

    Chan, Anthony W.S; Yang, Shang-Hsun

    2009-01-01

    Modeling human diseases using nonhuman primates including chimpanzee, rhesus, cynomolgus, marmoset and squirrel monkeys has been reported in the past decades. Due to the high similarity between nonhuman primates and humans, including genome constitution, cognitive behavioral functions, anatomical structure, metabolic, reproductive, and brain functions; nonhuman primates have played an important role in understanding physiological functions of the human body, clarifying the underlying mechanism of human diseases, and the development of novel treatments for human diseases. However, nonhuman primate research has been restricted to cognitive, behavioral, biochemical and pharmacological approaches of human diseases due to the limitation of gene transfer technology in nonhuman primates. The recent advancement in transgenic technology that has led to the generation of the first transgenic monkey in 2001 and a transgenic monkey model of Huntington's disease (HD) in 2008 has changed that focus. The creation of transgenic HD monkeys that replicate key pathological features of human HD patients further suggests the crucial role of nonhuman primates in the future development of biomedicine. These successes have opened the door to genetic manipulation in nonhuman primates and a new era in modeling human inherited genetic disorders. We focused on the procedures in creating transgenic Huntington's disease monkeys, but our work can be applied to transgenesis in other nonhuman primate species. PMID:19467335

  14. Design rules for efficient transgene expression in plants.

    PubMed

    Jackson, Mark A; Sternes, Peter R; Mudge, Stephen R; Graham, Michael W; Birch, Robert G

    2014-09-01

    Sustained expression of transgenes in specified developmental patterns is commonly needed in plant biotechnology, but obstructed by transgene silencing. Here, we present a set of gene design rules, tested on the silencing-susceptible beetle luc and bacterial ims genes, expressed in sugarcane. Designs tested independently or in combination included removal of rare codons, removal of RNA instability sequences, blocking of likely endogenous sRNA binding sites and randomization of non-rare codons. Stable transgene expression analyses, on multiple independent lines per construct, showed greatest improvement from the removal of RNA instability sequences, accompanied by greatly reduced transcript degradation evident in northern blot analysis. We provide a set of motifs that readily can be eliminated concurrently with rare codons and undesired structural features such as repeat sequences, using Gene Designer 2.0 software. These design rules yielded 935- and 5-fold increased expression in transgenic callus, relative to the native luc and ims sequences; and gave sustained expression under the control of sugarcane and heterologous promoters over several years in greenhouse and field trials. The rules can be applied easily with codon usage tables from any plant species, providing a simple and effective means to achieve sustained expression of otherwise silencing-prone transgenes in plants.

  15. Metabolic disruption identified in the Huntington's disease transgenic sheep model.

    PubMed

    Handley, Renee R; Reid, Suzanne J; Patassini, Stefano; Rudiger, Skye R; Obolonkin, Vladimir; McLaughlan, Clive J; Jacobsen, Jessie C; Gusella, James F; MacDonald, Marcy E; Waldvogel, Henry J; Bawden, C Simon; Faull, Richard L M; Snell, Russell G

    2016-02-11

    Huntington's disease (HD) is a dominantly inherited, progressive neurodegenerative disorder caused by a CAG repeat expansion within exon 1 of HTT, encoding huntingtin. There are no therapies that can delay the progression of this devastating disease. One feature of HD that may play a critical role in its pathogenesis is metabolic disruption. Consequently, we undertook a comparative study of metabolites in our transgenic sheep model of HD (OVT73). This model does not display overt symptoms of HD but has circadian rhythm alterations and molecular changes characteristic of the early phase disease. Quantitative metabolite profiles were generated from the motor cortex, hippocampus, cerebellum and liver tissue of 5 year old transgenic sheep and matched controls by gas chromatography-mass spectrometry. Differentially abundant metabolites were evident in the cerebellum and liver. There was striking tissue-specificity, with predominantly amino acids affected in the transgenic cerebellum and fatty acids in the transgenic liver, which together may indicate a hyper-metabolic state. Furthermore, there were more strong pair-wise correlations of metabolite abundance in transgenic than in wild-type cerebellum and liver, suggesting altered metabolic constraints. Together these differences indicate a metabolic disruption in the sheep model of HD and could provide insight into the presymptomatic human disease.

  16. Nutritional composition analysis of meat from human lactoferrin transgenic bulls.

    PubMed

    Zhao, Jie; Xu, Jianxiang; Wang, Jianwu; Li, Ning

    2013-01-01

    Transgenic technology has many potential advantages in food production. However, the transgenic technology process may influence the composition of food products derived from genetically engineered (GE) animals, which may be adverse to human health. Therefore, it is very important to research the compositions of GE animal products. Here, we analyzed the compositions of meat from the offspring of human lactoferrin (hLF) transgenic cows, which can express human lactoferrin proteins in their mammary gland. Six hLF transgenic bulls and three wide-type (WT) bulls, 10 months of age, were slaughtered for meat composition analysis. To determine the comparative health of hLF bulls for meat analysis, hematological analyses, organ/body weight analyses and pathology analyses were conducted. Results of the meat analysis show that there were no significant differences in the hematological parameters, organ/body weight ratios of hLF and WT bulls (P>0.05), and histopathological examination of the main organs of hLF bulls revealed no abnormalities. Nutrient parameters of meat compositions of hLF and WT bulls did not show any significant differences (P>0.05). All of these results suggest that the hLF transgene did not have an impact on the meat nutrient compositions of hLF bulls.

  17. Accelerated hepatocellular carcinoma development in CUL4B transgenic mice.

    PubMed

    Yuan, Jupeng; Jiang, Baichun; Zhang, Aizhen; Qian, Yanyan; Tan, Haining; Gao, Jiangang; Shao, Changshun; Gong, Yaoqin

    2015-06-20

    Cullin 4B (CUL4B) is a component of the Cullin 4B-Ring E3 ligase (CRL4B) complex that functions in proteolysis and in epigenetic regulation. CUL4B possesses tumor-promoting properties and is markedly upregulated in many types of human cancers. To determine the role of CUL4B in liver tumorigenesis, we generated transgenic mice that expressed human CUL4B in livers and other tissues and evaluated the development of spontaneous and chemically-induced hepatocellular carcinomas. We observed that CUL4B transgenic mice spontaneously developed liver tumors at a high incidence at old ages and exhibited enhanced DEN-induced hepatocarcinogenesis. There was a high proliferation rate in the livers of CUL4B transgenic mice that was accompanied by increased levels of Cdk1, Cdk4 and cyclin D1 and decreased level of p16. The transgenic mice also exhibited increased compensatory proliferation after DEN-induced liver injury, which was accompanied by activation of Akt, Erk, p38 and NF-κB. We also found that Prdx3 was downregulated and that DEN induced a higher level of reactive oxygen species in the livers of transgenic mice. Together, our results demonstrate a critical role of CUL4B in hepatocarcinogenesis in mice.

  18. Antibiotic multiresistance plasmid pRSB101 isolated from a wastewater treatment plant is related to plasmids residing in phytopathogenic bacteria and carries eight different resistance determinants including a multidrug transport system.

    PubMed

    Szczepanowski, Rafael; Krahn, Irene; Linke, Burkhard; Goesmann, Alexander; Pühler, Alfred; Schlüter, Andreas

    2004-11-01

    Ten different antibiotic resistance plasmids conferring high-level erythromycin resistance were isolated from an activated sludge bacterial community of a wastewater treatment plant by applying a transformation-based approach. One of these plasmids, designated pRSB101, mediates resistance to tetracycline, erythromycin, roxythromycin, sulfonamides, cephalosporins, spectinomycin, streptomycin, trimethoprim, nalidixic acid and low concentrations of norfloxacin. Plasmid pRSB101 was completely sequenced and annotated. Its size is 47 829 bp. Conserved synteny exists between the pRSB101 replication/partition (rep/par) module and the pXAC33-replicon from the phytopathogen Xanthomonas axonopodis pv. citri. The second pRSB101 backbone module encodes a three-Mob-protein type mobilization (mob) system with homology to that of IncQ-like plasmids. Plasmid pRSB101 is mobilizable with the help of the IncP-1alpha plasmid RP4 providing transfer functions in trans. A 20 kb resistance region on pRSB101 is located within an integron-containing Tn402-like transposon. The variable region of the class 1 integron carries the genes dhfr1 for a dihydrofolate reductase, aadA2 for a spectinomycin/streptomycin adenylyltransferase and bla(TLA-2) for a so far unknown Ambler class A extended spectrum beta-lactamase. The integron-specific 3'-segment (qacEDelta1-sul1-orf5Delta) is connected to a macrolide resistance operon consisting of the genes mph(A) (macrolide 2'-phosphotransferase I), mrx (hydrophobic protein of unknown function) and mphR(A) (regulatory protein). Finally, a putative mobile element with the tetracycline resistance genes tetA (tetracycline efflux pump) and tetR was identified upstream of the Tn402-specific transposase gene tniA. The second 'genetic load' region on pRSB101 harbours four distinct mobile genetic elements, another integron belonging to a new class and footprints of two more transposable elements. A tripartite multidrug (MDR) transporter consisting of an ATP

  19. Digital gene expression analysis of mature seeds of transgenic maize overexpressing Aspergillus niger phyA2 and its non-transgenic counterpart.

    PubMed

    Rao, Jun; Yang, Litao; Wang, Congmao; Zhang, Dabing; Shi, Jianxin

    2013-01-01

    The next generation sequencing technologies have been recently used for transcriptome analysis in many organisms because of the decreased sequencing cost and increased sequence output. In this study, we used digital gene expression (DGE) technique to compare the transcriptomic changes in mature seeds between transgenic maize overexpressing Aspergillus niger phyA2 and its non-transgenic counterpart. Deep sequencing of DGE libraries of the transgenic and its non-transgenic counterpart seeds generated 3,783,500 and 3,790,500 reads of 21-nucleotide, respectively, with frequencies spanning over four orders of magnitude. In transgenic maize, 53.97% of the unambiguous signature tags were mapped to the maize B73 reference genome, and 46.47% of genes were detected by at least two reads; in non-transgenic maize, the corresponding numbers were 51.38% and 47.39%. Compared with non-transgenic counterpart, about 12% of detected genes were differentially expressed in the transcriptome of transgenic maize seeds. Among these differentially expressed genes, there were 23 transcription factors in 14 families and no allergen genes. Pathway enrichment analysis revealed that 21 pathways were significantly affected by the transgenic event, in which the pathway involved in protein processing in endoplasmic reticulum was the most significantly affected. Results from this study indicated that both intended and unintended transcriptomic changes occurred in the transgenic maize, thus emphasizing the importance of transcriptome profiling in risk assessment of transgenic events.

  20. Molecular analysis, cytogenetics and fertility of introgression lines from transgenic wheat to Aegilops cylindrica host.

    PubMed

    Schoenenberger, Nicola; Guadagnuolo, Roberto; Savova-Bianchi, Dessislava; Küpfer, Philippe; Felber, François

    2006-12-01

    Natural hybridization and backcrossing between Aegilops cylindrica and Triticum aestivum can lead to introgression of wheat DNA into the wild species. Hybrids between Ae. cylindrica and wheat lines bearing herbicide resistance (bar), reporter (gus), fungal disease resistance (kp4), and increased insect tolerance (gna) transgenes were produced by pollination of emasculated Ae. cylindrica plants. F1 hybrids were backcrossed to Ae. cylindrica under open-pollination conditions, and first backcrosses were selfed using pollen bags. Female fertility of F1 ranged from 0.03 to 0.6%. Eighteen percent of the sown BC1s germinated and flowered. Chromosome numbers ranged from 30 to 84 and several of the plants bore wheat-specific sequence-characterized amplified regions (SCARs) and the bar gene. Self fertility in two BC1 plants was 0.16 and 5.21%, and the others were completely self-sterile. Among 19 BC1S1 individuals one plant was transgenic, had 43 chromosomes, contained the bar gene, and survived glufosinate treatments. The other BC1S1 plants had between 28 and 31 chromosomes, and several of them carried SCARs specific to wheat A and D genomes. Fertility of these plants was higher under open-pollination conditions than by selfing and did not necessarily correlate with even or euploid chromosome number. Some individuals having supernumerary wheat chromosomes recovered full fertility.

  1. BrUGE1 transgenic rice showed improved growth performance with enhanced drought tolerance.

    PubMed

    Abdula, Sailila E; Lee, Hye Jung; Kim, Joonki; Niño, Marjohn C; Jung, Yu-Jin; Cho, Young-Chan; Nou, Illsup; Kang, Kwon-Kyoo; Cho, Yong-Gu

    2016-03-01

    UDP-glucose 4-epimerase (UGE) catalyzes the reversible conversion of UDP-glucose to UDP-galactose. To understand the biological function of UGE from Brassica rapa, the gene BrUGE1 was cloned and introduced into the genome of wild type rice 'Gopum' using the Agrobacterium-mediated transformation method. Four lines which carried a single copy gene were selected and forwarded to T3 generation. Agronomic traits evaluation of the transgenic T3 lines (CB01, CB03, and CB06) under optimal field conditions revealed enriched biomass production particularly in panicle length, number of productive tillers, number of spikelets per panicle, and filled spikelets. These remarkably improved agronomic traits were ascribed to a higher photosynthetic rate complemented with higher CO2 assimilation. Transcripts of BrUGE1 in transgenic lines continuously accumulated at higher levels after the 20% PEG6000 treatment, implying its probable role in drought stress regulation. This was paralleled by rapid accumulation of soluble sugars which act as osmoprotectants, leading to delayed leaf rolling and drying. Our findings suggest the potential of BrUGE1 in improving rice growth performance under optimal and water deficit conditions.

  2. Abnormal B-cell function in HTLV-I-tax transgenic mice.

    PubMed

    Peebles, R S; Maliszewski, C R; Sato, T A; Hanley-Hyde, J; Maroulakou, I G; Hunziker, R; Schneck, J P; Green, J E

    1995-03-16

    Transgenic mice that carry the HTLV-I Tax gene develop an exocrinopathy with some similarities to Sjoegren's syndrome. Our experiments reveal that these mice have lymphadenopathy and splenomegaly composed primarily of B lymphocytes, as well as abnormal levels of secreted immunoglobulins. To gain insight into whether the lymphadenopathy manifested by these transgenic mice was the result of induction of cytokines by Tax, we utilized cell lines from these mice to study in vitro B-cell responses. Conditioned media (CM) derived from the cell lines caused B-cells to proliferate when a second signal, surface Ig cross-linking, was provided. The CM also caused a marked enhancement of IgM secretion by spleen cells or by purified B-cells treated with supplemental cytokines. The B-cell proliferative response and enhanced IgM secretion have not been attributed to a known cytokine. These results suggest that the CM from the cell lines contain a factor(s) involved in novel pathways of B-cell growth and differentiation that may participate in the pathologic development of autoimmune disease.

  3. Pollen-mediated gene flow from transgenic cotton under greenhouse conditions is dependent on different pollinators

    PubMed Central

    Yan, Shuo; Zhu, Jialin; Zhu, Weilong; Li, Zhen; Shelton, Anthony M.; Luo, Junyu; Cui, Jinjie; Zhang, Qingwen; Liu, Xiaoxia

    2015-01-01

    With the large-scale release of genetically modified (GM) crops, there are ecological concerns on transgene movement from GM crops to non-GM counterparts and wild relatives. In this research, we conducted greenhouse experiments to measure pollen-mediated gene flow (PGF) in the absence and presence of pollinators (Bombus ignitus, Apis mellifera and Pieris rapae) in one GM cotton (resistant to the insect Helicoverpa armigera and the herbicide glyphosate) and two non-GM lines (Shiyuan321 and Hai7124) during 2012 and 2013. Our results revealed that: (1) PGF varied depending on the pollinator species, and was highest with B. ignitus (10.83%) and lowest with P. rapae (2.71%); (2) PGF with B. ignitus depended on the distance between GM and non-GM cottons; (3) total PGF to Shiyuan321 (8.61%) was higher than to Hai7124 (4.10%). To confirm gene flow, we tested hybrids carrying transgenes for their resistance to glyphosate and H. armigera, and most hybrids showed strong resistance to the herbicide and insect. Our research confirmed that PGF depended on pollinator species, distance between plants and the receptor plant. PMID:26525573

  4. Longitudinal assessment of a transgenic animal model of tauopathy by FDG-PET imaging.

    PubMed

    de Cristóbal, Javier; García-García, Luis; Delgado, Mercedes; Pérez, Mar; Pozo, Miguel A; Medina, Miguel

    2014-01-01

    Abnormal levels and hyperphosphorylation of tau protein have been proposed as the underlying cause of a group of neurodegenerative disorders collectively known as 'tauopathies'. The detrimental consequence is the loss of affinity between this protein and the microtubules, increased production of fibrillary aggregates, and the accumulation of insoluble intracellular neurofibrillary tangles. A similar phenotype can be observed in various preclinical models, which have been generated to study the role of tau protein in neurodegenerative disorders. In this study, we have analyzed the brain metabolic activity in an animal model of tauopathy (tauVLW transgenic mice), which has been previously reported to mimic some of the phenotypic features of these disorders. By using a non-invasive technique, positron emission tomography (PET), a longitudinal non-clinical follow up study was carried out during most of the lifespan of these transgenic mice, from the youth to the senescence stages. The results obtained point out to an aging-dependent decrease in 18F-fluoro-deoxyglucose (FDG) uptake in the cerebral areas analyzed, which was already significant at the adult age, i.e., 11 months, and became much more prominent in the oldest animals (19 months old). This observation correlates well with the histopathological observation of neurodegeneration in brain areas where there is overexpression of tau protein.

  5. Use of transgenic seeds in Brazilian agriculture and concentration of agricultural production to large agribusinesses.

    PubMed

    Marinho, C D; Martins, F J O; Amaral Júnior, A T; Gonçalves, L S A; Amaral, S C S; de Mello, M P

    2012-07-19

    We identified the commercial releases of genetically modified organisms (GMOs) in Brazil, their characteristics, the types of genetic transformation used, and the companies responsible for the development of these GMOs, classifying them into two categories: private companies, subdivided into multinational and national, and public institutions. The data came from the data bank of the national registration of cultivars and the service of national protection of cultivars of the Ministry of Agriculture, Fishing and Supply (MAPA). This survey was carried out from 1998 to February 12, 2011. Until this date, 27 GMOs had been approved, including five for soybean, 15 for maize and seven for cotton cultivars. These GMOs have been used for the development of 766 cultivars, of which, 305 are soybean, 445 are maize, and 13 are cotton cultivars. The Monsato Company controls 73.2% of the transgenic cultivars certified by the MAPA; a partnership between Dow AgroSciences and DuPont accounts for 21.4%, and Syngenta controls 4.96%. Seed supply by these companies is almost a monopoly supported by law, giving no choice for producers and leading to the fast replacement of conventional cultivars by transgenic cultivars, which are expensive and exclude small producers from the market, since seeds cannot be kept for later use. This situation concentrates production in the hands of a few large national agribusiness entrepreneurs.

  6. Recombinant Human Myelin-Associated Glycoprotein Promoter Drives Selective AAV-Mediated Transgene Expression in Oligodendrocytes

    PubMed Central

    von Jonquieres, Georg; Fröhlich, Dominik; Klugmann, Claudia B.; Wen, Xin; Harasta, Anne E.; Ramkumar, Roshini; Spencer, Ziggy H. T.; Housley, Gary D.; Klugmann, Matthias

    2016-01-01

    Leukodystrophies are hereditary central white matter disorders caused by oligodendrocyte dysfunction. Recent clinical trials for some of these devastating neurological conditions have employed an ex vivo gene therapy approach that showed improved endpoints because cross-correction of affected myelin-forming cells occurred following secretion of therapeutic proteins by transduced autologous grafts. However, direct gene transfer to oligodendrocytes is required for the majority of leukodystrophies with underlying mutations in genes encoding non-secreted oligodendroglial proteins. Recombinant adeno-associated viral (AAV) vectors are versatile tools for gene transfer to the central nervous system (CNS) and proof-of-concept studies in rodents have shown that the use of cellular promoters is sufficient to target AAV-mediated transgene expression to glia. The potential of this strategy has not been exploited. The major caveat of the AAV system is its limited packaging capacity of ~5 kb, providing the rationale for identifying small yet selective recombinant promoters. Here, we characterize the human myelin associated glycoprotein (MAG) promoter for reliable targeting of AAV-mediated transgene expression to oligodendrocytes in vivo. A homology screen revealed highly conserved genomic regions among mammalian species upstream of the transcription start site. Recombinant AAV expression cassettes carrying the cDNA encoding enhanced green fluorescent protein (GFP) driven by truncated versions of the recombinant MAG promoter (2.2, 1.5 and 0.3 kb in size) were packaged as cy5 vectors and delivered into the dorsal striatum of mice. At 3 weeks post-injection, oligodendrocytes, neurons and astrocytes expressing the reporter were quantified by immunohistochemical staining. Our results revealed that both 2.2 and 1.5 kb MAG promoters targeted more than 95% of transgene expression to oligodendrocytes. Even the short 0.3 kb fragment conveyed high oligodendroglial specific transgene

  7. Recombinant Human Myelin-Associated Glycoprotein Promoter Drives Selective AAV-Mediated Transgene Expression in Oligodendrocytes.

    PubMed

    von Jonquieres, Georg; Fröhlich, Dominik; Klugmann, Claudia B; Wen, Xin; Harasta, Anne E; Ramkumar, Roshini; Spencer, Ziggy H T; Housley, Gary D; Klugmann, Matthias

    2016-01-01

    Leukodystrophies are hereditary central white matter disorders caused by oligodendrocyte dysfunction. Recent clinical trials for some of these devastating neurological conditions have employed an ex vivo gene therapy approach that showed improved endpoints because cross-correction of affected myelin-forming cells occurred following secretion of therapeutic proteins by transduced autologous grafts. However, direct gene transfer to oligodendrocytes is required for the majority of leukodystrophies with underlying mutations in genes encoding non-secreted oligodendroglial proteins. Recombinant adeno-associated viral (AAV) vectors are versatile tools for gene transfer to the central nervous system (CNS) and proof-of-concept studies in rodents have shown that the use of cellular promoters is sufficient to target AAV-mediated transgene expression to glia. The potential of this strategy has not been exploited. The major caveat of the AAV system is its limited packaging capacity of ~5 kb, providing the rationale for identifying small yet selective recombinant promoters. Here, we characterize the human myelin associated glycoprotein (MAG) promoter for reliable targeting of AAV-mediated transgene expression to oligodendrocytes in vivo. A homology screen revealed highly conserved genomic regions among mammalian species upstream of the transcription start site. Recombinant AAV expression cassettes carrying the cDNA encoding enhanced green fluorescent protein (GFP) driven by truncated versions of the recombinant MAG promoter (2.2, 1.5 and 0.3 kb in size) were packaged as cy5 vectors and delivered into the dorsal striatum of mice. At 3 weeks post-injection, oligodendrocytes, neurons and astrocytes expressing the reporter were quantified by immunohistochemical staining. Our results revealed that both 2.2 and 1.5 kb MAG promoters targeted more than 95% of transgene expression to oligodendrocytes. Even the short 0.3 kb fragment conveyed high oligodendroglial specific transgene

  8. Limited Fitness Advantages of Crop-Weed Hybrid Progeny Containing Insect-Resistant Transgenes (Bt/CpTI) in Transgenic Rice Field

    PubMed Central

    Yang, Xiao; Wang, Feng; Su, Jun; Lu, Bao-Rong

    2012-01-01

    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 F4 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. PMID:22815975

  9. Enhanced phytoremediation of volatile environmental pollutants with transgenic trees

    PubMed Central

    Doty, Sharon L.; James, C. Andrew; Moore, Allison L.; Vajzovic, Azra; Singleton, Glenda L.; Ma, Caiping; Khan, Zareen; Xin, Gang; Kang, Jun Won; Park, Jin Young; Meilan, Richard; Strauss, Steven H.; Wilkerson, Jasmine; Farin, Federico; Strand, Stuart E.

    2007-01-01

    Small, volatile hydrocarbons, including trichloroethylene, vinyl chloride, carbon tetrachloride, benzene, and chloroform, are common environmental pollutants that pose serious health effects. We have developed transgenic poplar (Populus tremula × Populus alba) plants with greatly increased rates of metabolism and removal of these pollutants through the overexpression of cytochrome P450 2E1, a key enzyme in the metabolism of a variety of halogenated compounds. The transgenic poplar plants exhibited increased removal rates of these pollutants from hydroponic solution. When the plants were exposed to gaseous trichloroethylene, chloroform, and benzene, they also demonstrated superior removal of the pollutants from the air. In view of their large size and extensive root systems, these transgenic poplars may provide the means to effectively remediate sites contaminated with a variety of pollutants at much faster rates and at lower costs than can be achieved with current conventional techniques. PMID:17940038

  10. Transgenic cells with increased plastoquinone levels and methods of use

    SciTech Connect

    Sayre, Richard T.; Subramanian, Sowmya; Cahoon, Edgar

    2016-12-27

    Disclosed herein are transgenic cells expressing a heterologous nucleic acid encoding a prephenate dehydrogenase (PDH) protein, a heterologous nucleic acid encoding a homogentisate solanesyl transferase (HST) protein, a heterologous nucleic acid encoding a deoxyxylulose phosphate synthase (DXS) protein, or a combination of two or more thereof. In particular examples, the disclosed transgenic cells have increased plastoquinone levels. Also disclosed are methods of increasing cell growth rates or production of biomass by cultivating transgenic cells expressing a heterologous nucleic acid encoding a PDH protein, a heterologous nucleic acid encoding an HST protein, a heterologous nucleic acid encoding a DXS protein, or a combination of two or more thereof under conditions sufficient to produce cell growth or biomass.

  11. Identification of transgenic foods using NIR spectroscopy: A review

    NASA Astrophysics Data System (ADS)

    Alishahi, A.; Farahmand, H.; Prieto, N.; Cozzolino, D.

    2010-01-01

    The utilization of chemometric methods in the quantitative and qualitative analysis of feeds, foods, medicine and so on has been accompanied with the great evolution in the progress and in the near infrared spectroscopy (NIRS). Hence, recently the application of NIR spectroscopy has extended on the context of genetics and transgenic products. The aim of this review was to investigate the application of NIR spectroscopy to identificate transgenic products and to compare it with the traditional methods. The results of copious researches showed that the application of NIRS technology was successful to distinguish transgenic foods and it has advantages such as fast, avoiding time-consuming, non-destructive and low cost in relation to the antecedent methods such as PCR and ELISA.

  12. Primary demyelination in transgenic mice expressing interferon-γ

    PubMed Central

    Mcgavern, Dorian B.; Rodriguez, Moses; Oldstone, Michael B.A.

    2017-01-01

    Ever since the use of interferon-γ to treat patients with multiple sclerosis resulted in enhanced disease, the role of IFN-γ in demyelinatlon has been under question. To address this issue directly, transgenic mice were generated that expressed the cDNA of murlne IFN-γ in the central nervous system by using an oligodendrocyte-specific promoter. Expression of the transgene occurred after 8 weeks of age, at which time the murlne immune and central nervous systems are both fully developed. Directly associated with transgene expression, primary demyelination occurred and was accompanied by clinical abnormalities consistent with CNS disorders. Additionally, multiple hallmarks of immune-mediated CNS disease were observed including upregulation of MHC molecules, gliosis and lymphocytlc infiltration. These results demonstrate a direct role for IFN-γ as an Inducer of CNS demyellnatlon leading to disease and provide new opportunities for dissecting the mechanism of demyelinatlon. PMID:9288735

  13. Transcriptional dysregulation in a transgenic model of Parkinson disease.

    PubMed

    Yacoubian, Talene A; Cantuti-Castelvetri, Ippolita; Bouzou, Bérengère; Asteris, Georgios; McLean, Pamela J; Hyman, Bradley T; Standaert, David G

    2008-03-01

    Alpha-synuclein has been implicated in Parkinson disease, yet the mechanism by which alpha-synuclein causes cell injury is not understood. Using a transgenic mouse model, we evaluated the effect of alpha-synuclein overexpression on gene expression in the substantia nigra. Nigral mRNA from wild type and alpha-synuclein transgenic mice was analyzed using Affymetrix gene arrays. At 3 months, before pathological changes are apparent, we observed modest alterations in gene expression. However, nearly 200 genes were altered in expression at 9 months, when degenerative changes are more apparent. Functional genomic analysis revealed that the genes altered at 9 months were predominantly involved in gene transcription. As in human Parkinson disease, gene expression changes in the transgenic model were also modulated by gender. These data demonstrate that alterations of gene expression are widespread in this animal model, and suggest that transcriptional dysregulation may be a disease mechanism that can be targeted therapeutically.

  14. [PCR detection of transgenic elements in feed raw material].

    PubMed

    Wang, Ying; Yu, Dao-Jian; Kang, Lin; Zhang, Gui-Ming; Jin, Xian-Zhong; Yang, Wei-Dong; Huang, Pei-Qing; Wu, Qiong; Chen, Zhi-Nan; Chu, Cheng-Cai; Cheng, Ying-Hui

    2002-05-01

    Based on the heterogeneous genes usually used in transgenic crops, the PCR technique was performed with primers derived from CaMV 35S promoter (35S-promoter,originated from cauliflower mosaic virus), NOS terminator (nopaline synthase-terminator,derived from Agrobacterium tumefaciens), EPSPS (5-enolpyruvylshikimate-3-phosphate synthase) gene, and CryIA(b) (delta-endotoxin,evolved from Bacillus thuringiensis subsp. kurstaki) gene to detect transgenic agents from feed raw materials of soybean dregs and corn gluten meal, respectively. Endogenous corn Zein (a protein extracted from corn gluten) gene, soybean Lectin (chitin-binding protein) gene and negative, positive control were applied for avoiding false results. The method established here has been successfully applied in detecting transgenic elements in imported feed raw material.

  15. Transgenic animal models of neurodegeneration based on human genetic studies

    PubMed Central

    Richie, Christopher T.; Hoffer, Barry J.; Airavaara, Mikko

    2011-01-01

    The identification of genes linked to neurodegenerative diseases such as Alzheimer's disease (AD), amyotrophic lateral sclerosis (ALS), Huntington's disease (HD) and Parkinson's disease (PD) has led to the development of animal models for studying mechanism and evaluating potential therapies. None of the transgenic models developed based on disease-associated genes have been able to fully recapitulate the behavioral and pathological features of the corresponding disease. However, there has been enormous progress made in identifying potential therapeutic targets and understanding some of the common mechanisms of neurodegeneration. In this review, we will discuss transgenic animal models for AD, ALS, HD and PD that are based on human genetic studies. All of the diseases discussed have active or complete clinical trials for experimental treatments that benefited from transgenic models of the disease. PMID:20931247

  16. National attitudes concerning gun carrying in the United States

    PubMed Central

    Hemenway, D; Azrael, D; Miller, M

    2001-01-01

    Objectives—To determine public attitudes in the United States concerning gun carrying. Setting—In the past 15 years, many state legislatures have passed laws making it easier for United States citizens to carry concealed firearms, not only on the street but into various locations, including churches and government buildings. Methods—National random digit dial telephone surveys conducted in 1996 and 1999 asked questions concerning the public's feelings of safety as more people in their community carry firearms, and whether, in the language of the question, respondents believe "regular" citizens should be allowed to carry guns into public or government buildings. Results—Americans feel less safe rather than more safe as more people in their community begin to carry guns. By margins of at least nine to one, Americans do not believe that "regular" citizens should be allowed to bring their guns into restaurants, college campuses, sports stadiums, bars, hospitals, or government buildings. Conclusions—The public believes that increased gun carrying by others reduces rather than increases their safety. Overwhelmingly, the public believes that in many venues gun carrying should be prohibited. PMID:11770652

  17. Establishment of a novel, eco-friendly transgenic pig model using porcine pancreatic amylase promoter-driven fungal cellulase transgenes.

    PubMed

    Lin, Y S; Yang, C C; Hsu, C C; Hsu, J T; Wu, S C; Lin, C J; Cheng, W T K

    2015-02-01

    Competition between humans and livestock for cereal and legume grains makes it challenging to provide economical feeds to livestock animals. Recent increases in corn and soybean prices have had a significant impact on the cost of feed for pig producers. The utilization of byproducts and alternative ingredients in pig diets has the potential to reduce feed costs. Moreover, unlike ruminants, pigs have limited ability to utilize diets with high fiber content because they lack endogenous enzymes capable of breaking down nonstarch polysaccharides into simple sugars. Here, we investigated the feasibility of a transgenic strategy in which expression of the fungal cellulase transgene was driven by the porcine pancreatic amylase promoter in pigs. A 2,488 bp 5'-flanking region of the porcine pancreatic amylase gene was cloned by the genomic walking technique, and its structural features were characterized. Using GFP as a reporter, we found that this region contained promoter activity and had the potential to control heterologous gene expression. Transgenic pigs were generated by pronuclear microinjection. Founders and offspring were identified by PCR and Southern blot analyses. Cellulase mRNA and protein showed tissue-specific expression in the pancreas of F1 generation pigs. Cellulolytic enzyme activity was also identified in the pancreas of transgenic pigs. These results demonstrated the establishment of a tissue-specific promoter of the porcine pancreatic amylase gene. Transgenic pigs expressing exogenous cellulase may represent a way to increase the intake of low-cost, fiber-rich feeds.

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

    PubMed Central

    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

  19. Predator odor exposure increases food-carrying behavior in rats.

    PubMed

    Wernecke, Kerstin E A; Brüggemann, Judith; Fendt, Markus

    2016-02-01

    To cover their energy demands, prey animals are forced to search for food. However, during foraging they also expose themselves to the risk of becoming the prey of predators. Consequently, in order to increase their fitness foraging animals have to trade-off efficiency of foraging against the avoidance of predation risk. For example, the decision on whether a found food piece should be eaten at the food source or whether it should be carried to a protective site such as the nest (food-carrying behavior), is strongly dependent on different incentive factors (e.g., hunger level, food size, distance to the nest). It has been shown that food-carrying behavior increases the more risky the foraging situation becomes. Since predator odors are clearly fear-inducing in rats, we ask here whether the detection of predator odors in close proximity to the food source modulates food-carrying behavior. In the present study, the food-carrying behavior of rats for six different food pellet sizes was measured in a "low risk" and a "high risk" testing condition by presenting water or a fox urine sample, respectively, next to the food source. For both testing conditions, food-carrying behavior of rats increased with increasing food pellet weight. Importantly, the proportion of food-carrying rats was significantly higher during exposure to fox urine ("high risk") than when rats were tested with the water control ("low risk"). Taken together, these results demonstrate that food-carrying behavior of rats is increased by the detection of a predator odor. Our data also support the idea that such food-carrying behavior can be considered as a pre-encounter defensive response.

  20. Global change and carrying capacity: Implications for life on Earth

    NASA Technical Reports Server (NTRS)

    Ehrlich, Paul R.; Daily, Gretchen C.; Ehrlich, Anne H.; Matson, Pamela; Vitousek, Peter

    1989-01-01

    Determining the long-term number of people that the planet can support without irreversibly reducing its ability to support people in the future, i.e., the carrying capacity of the Earth, is an exceedingly complex problem. About all that is known for certain is that, with present and foreseeable technologies, the human population has already exceeded the capacity. The reduction in carrying capacity that can be expected to result from direct human impacts on resources and the environment and from our indirect impacts of the climate system is discussed. Global warming and modeling global change and food security are also discussed with respect to carrying capacity.