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Sample records for carrying mobilizable transgenes

  1. Plant transformation by coinoculation with a disarmed Agrobacterium tumefaciens strain and an Escherichia coli strain carrying mobilizable transgenes.

    PubMed

    Pappas, Katherine M; Winans, Stephen C

    2003-11-01

    Transformation of Nicotiana tabacum leaf explants was attempted with Escherichia coli as a DNA donor either alone or in combination with Agrobacterium tumefaciens. We constructed E. coli donor strains harboring either the promiscuous IncP-type or IncN-type conjugal transfer system and second plasmids containing the respective origins of transfer and plant-selectable markers. Neither of these conjugation systems was able to stably transform plant cells at detectable levels, even when VirE2 was expressed in the donor cells. However, when an E. coli strain expressing the IncN-type conjugation system was coinoculated with a disarmed A. tumefaciens strain, plant tumors arose at high frequencies. This was caused by a two-step process in which the IncN transfer system mobilized the entire shuttle plasmid from E. coli to the disarmed A. tumefaciens strain, which in turn processed the T-DNA and transferred it to recipient plant cells. The mobilizable plasmid does not require a broad-host-range replication origin for this process to occur, thus reducing its size and genetic complexity. Tumorigenesis efficiency was further enhanced by incubation of the bacterial strains on medium optimized for bacterial conjugation prior to inoculation of leaf explants. These techniques circumvent the need to construct A. tumefaciens strains containing binary vectors and could simplify the creation of transgenic plants. PMID:14602634

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

    PubMed

    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.

  3. 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

  4. Allelic exclusion in transgenic mice carrying mutant human IgM genes

    PubMed Central

    1988-01-01

    Expression of the membrane-bound version of the human mu chain in transgenic mice results in the allelic exclusion of endogenous mouse Ig heavy chain genes (6). The secreted version of the human Ig transgene has no such effect. F1 hybrid animals that carry transgenes for both secreted and membrane-bound human mu chains produce both forms of the human heavy chain while strongly suppressing endogenous mouse mu expression. The simultaneous expression of the two rearranged transgenes in primary B cells suggests that allelic exclusion operates before the formation of a second functionally rearranged heavy chain gene in vivo. PMID:3133444

  5. 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

  6. Mobilizable RDF/d-RDF burning program

    SciTech Connect

    Niemann, K.; Campbell, J.

    1982-03-01

    The Mobilizable RDF/d-RDF Burning Program was conceived to promote the utilization of refuse-derived fuels (RDF) as a supplement to existing fossil fuel sources in industrial-sized boilers. The program explores the design, development, and eventual construction of densified-RDF (d-RDF) for use in boiler combustion testing as a supplement to stoker coal or wood wastes. The equipment would be mounted on trailers and assembled and operated at preselected sites throughout the country where approximately 750 tons of RDF would be produced and test burned in a local boiler. The equipment, to include a transportable RDF boiler metering and feed system, would then be moved and operated at two to three test sites annually. The program is intended to encourage the construction of permanent resource recovery facilities by involving local waste handling groups in operating the equipment and producing fuel, and potential local fuel users in testing the fuel in their boilers. The Mobilizable Program was developed from two separate tasks. The first task developed the concept behind the program and defined its operational and organizational structure. The second task, a follow-up to the first, was intended principally to finalize test locations, develop equipment designs and specifications, and formalize a management program. This report summarizes the principal findings of both tasks. It identifies the criteria used to identify test locations, outlines the program's management structure, presents design and performance specifications for both the fuel production equipment and boiler fuel feed systems, and provides a detailed evaluation of the parameters involved in burning RDF in industrial-sized boilers. Final conclusions and recommendations identify problem areas encountered in the program, and discuss possible future directions for such a program.

  7. Generation and Characterization of a Transgenic Pig Carrying a DsRed-Monomer Reporter Gene

    PubMed Central

    Wu, Mei-Han; Yang, Cho-Chen; Lin, Yu-Sheng; Cheng, Winston Teng-Kui; Wu, Shinn-Chih; Lin, Yao-Ping

    2014-01-01

    Background Pigs are an optimal animal for conducting biomedical research because of their anatomical and physiological resemblance to humans. In contrast to the abundant resources available in the study of mice, few fluorescent protein-harboring porcine models are available for preclinical studies. In this paper, we report the successful generation and characterization of a transgenic DsRed-Monomer porcine model. Methods The transgene comprised a CMV enhancer/chicken-beta actin promoter and DsRed monomeric cDNA. Transgenic pigs were produced by using pronuclear microinjection. PCR and Southern blot analyses were applied for identification of the transgene. Histology, blood examinations and computed tomography were performed to study the health conditions. The pig amniotic fluid progenitor/stem cells were also isolated to examine the existence of red fluorescence and differentiation ability. Results Transgenic pigs were successfully generated and transmitted to offspring at a germ-line transmission rate of 43.59% (17/39). Ubiquitous expression of red fluorescence was detected in the brain, eye, tongue, heart, lung, liver, pancreas, spleen, stomach, small intestine, large intestine, kidney, testis, and muscle; this was confirmed by histology and western blot analyses. In addition, we confirmed the differentiation potential of amniotic fluid progenitor stem cells isolated from the transgenic pig. Conclusions This red fluorescent pig can serve as a host for other fluorescent-labeled cells in order to study cell-microenvironment interactions, and can provide optimal red-fluorescent-labeled cells and tissues for research in developmental biology, regenerative medicine, and xenotransplantation. PMID:25187950

  8. Lentiviral vectors carrying enhancer elements of Hb9 promoter drive selective transgene expression in mouse spinal cord motor neurons.

    PubMed

    Peviani, Marco; Kurosaki, Mami; Terao, Mineko; Lidonnici, Dario; Gensano, Francesco; Battaglia, Elisa; Tortarolo, Massimo; Piva, Roberto; Bendotti, Caterina

    2012-03-30

    Recombinant lentiviral vectors (rLVs) have emerged as versatile tools for gene delivery applications due to a number of favorable features, such as the possibility to maintain long-term transgene expression, the flexibility in the design of the expression cassettes and recent improvements in their biosafety profile. Since rLVs are able to infect multiple cell types including post-mitotic cells such as neurons and skeletal muscle cells, several studies have been exploring their application for the study and cure of neurodegenerative diseases. In particular, the introduction of rLVs carrying cell-type specific promoters could restrict the transgene expression either to neuronal or glial cells, thus helping to better dissect in vivo the role played by these cell populations in several neurodegenerative processes. In this study we developed rLVs carrying motor neuron specific regulatory sequences derived from the promoter of homeobox gene Hb9, and demonstrated that these constructs can represent a suitable platform for selective gene-targeting of murine spinal cord motor neurons, in vivo. This tool could be instrumental in the dissection of the molecular mechanisms involved in the selective degeneration of motor neurons occurring in Motor Neuron Diseases.

  9. 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.

  10. Hyperplasia and tumours in lung, breast and other tissues in mice carrying a RAR beta 4-like transgene.

    PubMed

    Bérard, J; Gaboury, L; Landers, M; De Repentigny, Y; Houle, B; Kothary, R; Bradley, W E

    1994-12-01

    Transgenic mice were generated which express a truncated nuclear retinoic acid receptor beta (RAR beta), closely resembling the natural isoform RAR beta 4, under the control of the MMTV promoter. The transgene was expressed in salivary gland, testis, lung and mammary tissue in two different lines. At approximately 11-14 months virtually all the transgenic mice showed hyperplasia of the lung alveolar epithelium with an excess of type II pneumocytes. Hyperplasia of the mammary alveoli and terminal ducts was also seen in some females. Salivary glands and some sebaceous glands were hyperplastic in most male transgenic mice, but only rarely in females or in non-transgenics. Primary benign and malignant tumours were more numerous in transgenic mice than in controls, with a total of 23 in 43 mice versus two in 33 non-transgenic animals. Treatment with dexamethasone to increase transgene expression resulted in exaggerated versions of the above phenotypes. Overexpression of RAR beta 4 therefore appears to predispose various tissues to hyperplasia and neoplasia, and this by contrast to the RAR beta 2 isoform, which has tumour suppressor activity. A survey of ratios of RAR beta 4:RAR beta 2 expression in human lung tumour cell lines showed an increase compared with normal lung tissue, suggesting that RAR beta 4 may play a similar role in human tumorigenesis.

  11. Adaptive regulation of intestinal thiamin uptake: molecular mechanism using wild-type and transgenic mice carrying hTHTR-1 and -2 promoters.

    PubMed

    Reidling, Jack C; Said, Hamid M

    2005-06-01

    Thiamin participates in metabolic pathways contributing to normal cellular functions, growth, and development. The molecular mechanism of the human intestinal thiamin absorption process involves the thiamin transporters-1 (hTHTR-1) and -2 (hTHTR-2), products of the SLC19A2 and SLC19A3 genes. Little is known about adaptive regulation of the intestinal thiamin uptake process or the molecular mechanism(s) involved during thiamin deficiency. In these studies, we addressed these issues using wild-type mice and transgenic animals carrying the promoters of the hTHTR-1 and -2. We show that, in thiamin deficiency, a significant and specific upregulation in intestinal carrier-mediated thiamin uptake occurs and that this increase is associated with an induction in protein and mRNA levels of mTHTR-2 but not mTHTR-1; in addition, an increase in the activity of the SLC19A3, but not the SLC19A2, promoter was observed in the intestine of transgenic mice. Similar findings were detected in the kidney; however, expression of both thiamin transporters and activity of both human promoters were upregulated in this organ in thiamin deficiency. We also examined the effect of thiamin deficiency on the level of expression of mTHTR-1 and mTHTR-2 messages and activity of the human promoters in the heart and brain of transgenic mice and found an increase in mTHTR-1 mRNA and a rise in activity of the SLC19A2 promoter in thiamin-deficient mice. These results show that the intestinal and renal thiamin uptake processes are adaptively upregulated during dietary thiamin deficiency, that expression of mTHTR-1 and mTHTR-2 is regulated in a tissue-specific manner, and that this upregulation is mediated via transcriptional regulatory mechanism(s).

  12. 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.

  13. 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

  14. The Tsukuba hypertensive mouse (transgenic mouse carrying human genes for both renin and angiotensinogen) as a model of human malignant hypertension: development of lesions and morphometric analysis.

    PubMed

    Shimokama, T; Haraoka, S; Horiguchi, H; Sugiyama, F; Murakami, K; Watanabe, T

    1998-02-01

    The renin-angiotensin system has a pivotal role in hypertension. The Tsukuba hypertensive mouse (THM; a transgenic mouse carrying human genes for both renin and angiotensinogen) was generated to allow further examination of the renin-angiotensin system in a variety of pathologic conditions. We evaluated the development of renal lesions in these mice and in controls by morphometric, immunohistochemical and ultrastructural methods. Blood pressure was significantly higher in THM than in control mice; 1 year after birth, it was approximately 40 mmHg higher. The kidney-to-body weight ratio was also higher in THM than in control. Morphometrical analysis revealed that the glomerular sclerosis index was significantly elevated in THM with 10% of the glomeruli sclerotic at 18 months. The grade of vascular lesion and the frequency of fibronoid arteritis of the kidney exhibited the same tendency as the glomerular sclerosis index. Murine renin was located exclusively in the juxtaglomerular apparatus, whereas human renin was expressed not only in the juxtaglomerular apparatus, but also in periarteriolar smooth muscle cells and in mesangial and epithelial cells of the glomeruli. Light and electron microscopy revealed significant fibrinoid arteritis of the kidney in THM and also "onion skinning", both pathognomonic for malignant nephrosclerosis. THM may be an excellent model of human malignant hypertension. PMID:9504863

  15. 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. PMID:25456072

  16. Induction of phase II enzymes and hsp70 genes by copper sulfate through the electrophile-responsive element (EpRE): insights obtained from a transgenic zebrafish model carrying an orthologous EpRE sequence of mammalian origin.

    PubMed

    Almeida, Daniela Volcan; Nornberg, Bruna Félix da Silva; Geracitano, Laura A; Barros, Daniela Martí; Monserrat, José Maria; Marins, Luis Fernando

    2010-09-01

    We have evaluated the homology of the electrophile-responsive element (EpRE) core sequence, a binding site for the Nrf2 transcription factor, in the proximal promoters of the mouse and zebrafish glutathione-S-transferase (gst), glutamate cysteine ligase catalytic subunit (gclc) and heat shock protein 70 (hsp70) genes. The EpRE sites identified for both species in the three analyzed genes showed a high similarity with the putative EpRE core sequence. We also produced a transgenic zebrafish model carrying a transgene comprised of the luciferase (luc) reporter gene under transcriptional control of a mouse EpRE sequence. This transgenic model was exposed to copper sulfate, and the reporter gene was significantly activated. The endogenous gst, gclc and hsp70 zebrafish genes were analyzed in the EpRE-Luc transgenic zebrafish and showed an expression pattern similar to that of the reporter transgene used. Our results demonstrate that EpRE is conserved between mouse and zebrafish for detoxification-related genes and that the development of genetically modified models using this responsive element to drive the expression of reporter genes can be an important tool in understanding the action mechanism of aquatic pollutants. PMID:19116768

  17. 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

  18. 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

    2014-01-01

    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.

  19. 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.

  20. Characterization of the integrase of NBU1, a Bacteroides mobilizable transposon.

    PubMed

    Rajeev, Lara; Salyers, Abigail A; Gardner, Jeffrey F

    2006-08-01

    NBU1 is a Bacteroides mobilizable transposon (MTn) that is integrated within the host chromosome and requires CTnDOT functions for its excision and transfer into a new host. The NBU1 integrase IntN1 has been classified as a tyrosine recombinase based on the presence of conserved residues. We created alanine mutants of the residues R291, K314, H393, R396, H419 and the conserved substitution Y429F and tested them for integration efficiency. The results suggest that these residues in IntN1 are important for integration, and Y429 could be the catalytic nucleophile. We employed suicide substrates and partially purified IntN1 to determine the positions of IntN1 cleavage within the 14 bp common core region that is identical in both NBU1 att sites. We show that IntN1 makes 7 bp staggered cuts on the top and bottom strands. From previous mutational analysis of the att sites, we show that two specific mutations near the site of bottom strand cleavage within this 7 bp region increased integration, and mutations of the two bases near top strand cleavage site had no effect on integration. These results indicate that IntN1 lacks the strict requirement for homology between the recombining sites seen with other tyrosine recombinases. We also show that phosphorothioate substitutions at the cleavage site and 1 bp downstream inhibited cleavage by IntN1. This differs from other studied tyrosine recombinases where inhibition occurs by substitutions at the cleavage site only.

  1. Construction of stably maintained non-mobilizable derivatives of RSF1010 lacking all known elements essential for mobilization

    PubMed Central

    Katashkina, Joanna I; Kuvaeva, Tatiana M; Andreeva, Irina G; Skorokhodova, Alexandra Yu; Biryukova, Irina V; Tokmakova, Irina L; Golubeva, Lubov I; Mashko, Sergey V

    2007-01-01

    Background RSF1010 is a well-studied broad-host-range plasmid able to be mobilized to different bacteria and plants. RSF1010-derived plasmid vectors are widely used in both basic research and industrial applications. In the latter case, exploiting of mobilizable plasmids or even the plasmids possessing negligible mobilization frequency, but containing DNA fragments that could promote conjugal transfer, is undesirable because of biosafety considerations. Previously, several mutations significantly decreasing efficiency of RSF1010 mobilization have been selected. Nevertheless, construction of the RSF1010 derivative lacking all known loci involved in the conjugal transfer has not been reported yet. Results Novel non-mobilizable derivatives of RSF1010 lacking all known DNA sequences involved in the mobilization process have been obtained due to the exploiting of λRed-driven recombination between the plasmid and a constructed in vitro linear DNA fragment. To provide auto-regulated transcription of the essential replication gene, repB, the plasmid loci oriT, mobC and mobA were substituted by the DNA fragment containing PlacUV5→lacI. Mobilization of the obtained RSFmob plasmid was not detected in standard tests. The derivative of RSFmob with increased copy number has been obtained after lacI elimination. High stability of both constructed plasmids has been demonstrated in Escherichia coli and Pantoea ananatis. Design of RSFmob allows easy substitution of PlacUV5 by any desirable promoter for construction of novel derivatives with changed copy number or host range. Conclusion Novel non-mobilizable derivatives of RSF1010 lacking all known DNA sequences involved in the mobilization process and stably maintained at least in E. coli and P. ananatis have been constructed. The obtained plasmids became the progenitors of new cloning vectors answering all biosafety requirements of genetically modified organisms used in scale-up production. PMID:18028554

  2. Transgenic mice containing a 248-kb yeast artificial chromosome carrying the human beta-globin locus display proper developmental control of human globin genes.

    PubMed Central

    Peterson, K R; Clegg, C H; Huxley, C; Josephson, B M; Haugen, H S; Furukawa, T; Stamatoyannopoulos, G

    1993-01-01

    Transgenic mice were generated using a purified 248-kb yeast artificial chromosome (YAC) bearing an intact 82-kb human beta-globin locus and 148 kb of flanking sequence. Seventeen of 148 F0 pups were transgenic. RNase protection analysis of RNA isolated from the blood of 13 gamma- and beta-globin-positive founders showed that only the human beta-globin gene was expressed in the adult founders. Studies of F1 and F2 fetuses demonstrated that the genes of the beta-locus YAC displayed the proper developmental switches in beta-like globin gene expression. Expression of epsilon- and gamma-globin, but not beta-globin, was observed in the yolk sac, there was only minor gamma and mostly beta expression in the 14-day liver, and only beta mRNA in the blood of the adult animals. Structural data showed that the locus was intact. These results indicate that it is now possible to dissect regulatory mechanisms within the context of an entire locus in vivo by using the ability to perform mutagenesis efficiently in yeast via homologous recombination, followed by purification of the altered YAC and its introduction into mice. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 PMID:8356061

  3. Assays of polychlorinated biphenyl congeners and co-contaminated heavy metals in the transgenic Arabidopsis plants carrying the recombinant guinea pig aryl hydrocarbon receptor-mediated β-glucuronidase reporter gene expression system.

    PubMed

    Shimazu, Sayuri; Ohta, Masaya; Ohkawa, Hideo; Ashida, Hitoshi

    2012-01-01

    The transgenic Arabidopsis plant XgD2V11-6 carrying the recombinant guinea pig (g) aryl hydrocarbon receptor (AhR)-mediated β-glucuronidase (GUS) reporter gene expression system was examined for assay of polychlorinated biphenyl (PCB) congeners and co-contaminated heavy metals. When the transgenic Arabidopsis plants were treated with PCB126 (toxic equivalency factor; TEF: 0.1) and PCB169 (TEF: 0.03), the GUS activity of the whole plants was increased significantly. After treatment with PCB80 (TEF: 0), the GUS activity was nearly the same level as that treated with 0.1% dimethylsulfoxide (DMSO) as a vehicle control. After exposure to a 1:1 mixture of PCB126 and PCB169, the GUS activity was increased additively. However, after exposure to a mixture of PCB126 and PCB80, the GUS activity was lower than that of the treatment with PCB126 alone. Thus, PCB80 seemed to be an antagonist towards AhR. When the transgenic plants were treated with each of the heavy metals Fe, Cu, Zn, Cd and Pb together with PCB126, Cd and Pb increased the PCB126-induced GUS activity. On the other hand, Fe, Cu and Zn did not affect the PCB126-induced GUS activity. In the presence of the biosurfactant mannosylerythritol lipid-B (MEL-B) and the carrier protein bovine serum albumin (BSA), the PCB126-induced GUS activity was increased, but the Cd-assisted PCB126-induced GUS activity was not affected. Thus, MEL-B and BSA seemed to increase uptake and transport of PCB126, respectively. PMID:22938576

  4. 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.

  5. Augmented oxygen-mediated transcriptional activation of cytochrome P450 (CYP)1A expression and increased susceptibilities to hyperoxic lung injury in transgenic mice carrying the human CYP1A1 or mouse 1A2 promoter in vivo.

    PubMed

    Jiang, Weiwu; Couroucli, Xanthi I; Wang, Lihua; Barrios, Roberto; Moorthy, Bhagavatula

    2011-04-01

    Supplemental oxygen administration is frequently administered to pre-term and term infants having pulmonary insufficiency. However, hyperoxia contributes to the development of bronchopulmonary dysplasia (BPD) in premature infants. Cytochrome P450 (CYP)A enzymes have been implicated in hyperoxic lung injury. In this study, we tested the hypothesis that hyperoxia induces CYP1A1 and 1A2 enzymes by transcriptional activation of the corresponding promoters in vivo, and transgenic mice expressing the human CYP1A1 or the mouse 1A2 promoter would be more susceptible to hyperoxic lung injury than wild type (WT) mice. Adult WT (CD-1) (12week-old) mice, transgenic mice carrying a 10kb human CYP1A1 promoter and the luciferase (luc) reporter gene (CYP1A1-luc), or mice expressing the mouse CYP1A2 promoter (CYP1A2-luc) were maintained in room air or exposed to hyperoxia for 24-72h. Hyperoxia exposure of CYP1A1-luc mice for 24 and 48h resulted in 2.5- and 1.25-fold increases, respectively, in signal intensities, compared to room air controls. By 72h, the induction had declined to control levels. CYP1A2-luc mice also showed enhanced luc expression after 24-48h, albeit to a lesser extent than those expressing the CYP1A1 promoter. Also, these mice showed decreased levels of endogenous CYP1A1 and 1A2 expression after prolonged hyperoxia, and were also more susceptible to lung injury than similarly exposed WT mice, with CYP1A2-luc mice showing the greatest injury. Our results support the hypothesis that hyperoxia induces CYP1A enzymes by transcriptional activation of its corresponding promoters, and that decreased endogenous expression of these enzymes contribute to the increased susceptibilities to hyperoxic lung injury in the transgenic animals. In summary, this is the first report providing direct evidence of hyperoxia-mediated induction of CYP1A1 and CYP1A2 expression in vivo by mechanisms entailing transcriptional activation of the corresponding promoters, a phenomenon that has

  6. 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

    2016-01-01

    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

  7. 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

  8. Cryopreservation of Xenopus transgenic lines.

    PubMed

    Buchholz, Daniel R; Fu, Liezhen; Shi, Yun-Bo

    2004-01-01

    Xenopus laevis has been widely used for molecular, cellular, and developmental studies. With the development of the sperm-mediated transgenic method, it is now possible to study gene function during vertebrate development by using this popular model. On the other hand, like other animal species, it is labor intensive, and the maintenance of transgenic lines is expensive. In this article, we investigated the possibility of using sperm-cryopreservation as a means to preserve transgenic frog lines. We demonstrated that cryopreserved sperms are viable but not fertile under our in vitro fertilization (IVF) conditions. However, by microinjecting cryopreserved sperm nuclei, we successfully regenerated a transgenic line carrying a double promoter transgene construct, where the marker gene encoding the green fluorescent protein (GFP) is driven by the gamma-crystallin gene promoter and a gene of interest, encoding a fusion protein of GFP with the matrix metalloproteinase stromelysin-3 (ST3-GFP), is driven by a heat shock-inducible promoter. We demonstrated the functional transmission of the ST3-GFP transgene by analyzing the phenotype of the F1 animals after heat-shock to induce its expression. Our method thus provides an inexpensive means to preserve transgenic frog lines and a convenient way for distribution of transgenic lines. Furthermore, the ease with which to microinject nuclei compared to the technically demanding transgenesis procedure with variable outcome should facilitate more laboratories to use transgenic Xenopus laevis for functional studies in vivo. Mol. Reprod. Dev. 67: 65-69, 2004.

  9. [The safety and usefulness of transgenic plants].

    PubMed

    Ondrej, M; Drobník, M

    1997-05-29

    Transgenic crop plants, used in food and feed production, carry different beneficial transgenes, mostly for resistance to pests, herbicides and diseases. All new transgenic plant varieties, the genes they carry and their products have been thoroughly tested before released for agriculture and even more for marketing. Genetically modified organisms carry the same risk as any other organism. Food derived from genetically modified organisms due to legal regulation is most controlled and therefore most safe food ever placed on the market. In future, transgenic plants offer many new possibilities also for medical use, like plant vaccines, antibiotics and rare proteins of clinical importance produced by plants.

  10. 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.…

  11. 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

  12. [Transgenic plants].

    PubMed

    Blum, H E

    2002-12-01

    Advances in molecular genetics and recombinant DNA technology have revolutionized our understanding of the pathogenesis as well as the diagnosis, therapy and prevention of human diseases. Similar developments characterize plant biotechnology with the production of plant derived biomedical as well as health products. Apart from the fundamentals of molecular plant genetics, the production of transgenic plants as well as the clinical relevance, benefits, limitations and potential problems of plant biotechnology will be reviewed in some detail. It is a particular challenge to physicians in an increasingly informed environment to be informed about the new developments in molecular biology and recombinant DNA technology and to have a qualified opinion about their clinical relevance.

  13. Transgenic mouse offspring generated by ROSI

    PubMed Central

    MOREIRA, Pedro; PÉREZ-CEREZALES, Serafín; LAGUNA, Ricardo; FERNÁNDEZ-GONZALEZ, Raúl; SANJUANBENITO, Belén Pintado; GUTIÉRREZ-ADÁN, Alfonso

    2015-01-01

    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. PMID:26498042

  14. Transgenic hepatocarcinogenesis in the rat.

    PubMed Central

    Hully, J. R.; Su, Y.; Lohse, J. K.; Griep, A. E.; Sattler, C. A.; Haas, M. J.; Dragan, Y.; Peterson, J.; Neveu, M.; Pitot, H. C.

    1994-01-01

    Although transgenic hepatocarcinogenesis has been accomplished in the mouse with a number of genetic constructs targeting the oncogene to expression primarily in the liver, no example of this process has yet been developed in the rat. Because our understanding of the multistage nature of hepatocarcinogenesis is most advanced in the rat, we have developed a strain of transgenic rats carrying the promoter-enhancer sequences of the mouse albumin gene linked 5' to the simian virus-40 T antigen gene. A line of transgenic rats bearing this transgene has been developed from a single founder female. Five to six copies of the transgene, possibly in tandem, occur within the genome of the transgenic animals, which are maintained by heterozygous matings. Livers of transgenic animals are histologically normal after weaning; at 2 months of age, small foci of vacuolated cells appear in this organ. By 4 months of age, all animals exhibit focal lesions and nodules consisting primarily of small basophilic cells, many of which exhibit considerable cytoplasmic vacuolization. Mating of animals each bearing the transgene results in rats with a demyelinating condition that develops acutely in pregnant females and more chronically in males. Ultrastructural studies of these cells indicate that the vacuoles contain substantial amounts of glycogen, with the cells resembling hepatoblasts. Malignant neoplasms with both a glandular and a hepatoblastoma/hepatocellular carcinoma pattern arise from the nodules. Enzyme and immunohistochemical studies of all lesions reveal many similarities in gene expression to comparable lesions in rats subjected to chemically induced hepatocarcinogenesis, with certain exceptions. The placental form of glutathione-S-transferase is absent from all lesions in the transgenic animal, as is the expression of connexin 32. A significant number of lesions express serum albumin, and many, but not all, exhibit the T antigen. Lesions expressing the T antigen also contain

  15. Transgenic resistance.

    PubMed

    Cillo, Fabrizio; Palukaitis, Peter

    2014-01-01

    Transgenic resistance to plant viruses is an important technology for control of plant virus infection, which has been demonstrated for many model systems, as well as for the most important plant viruses, in terms of the costs of crop losses to disease, and also for many other plant viruses infecting various fruits and vegetables. Different approaches have been used over the last 28 years to confer resistance, to ascertain whether particular genes or RNAs are more efficient at generating resistance, and to take advantage of advances in the biology of RNA interference to generate more efficient and environmentally safer, novel "resistance genes." The approaches used have been based on expression of various viral proteins (mostly capsid protein but also replicase proteins, movement proteins, and to a much lesser extent, other viral proteins), RNAs [sense RNAs (translatable or not), antisense RNAs, satellite RNAs, defective-interfering RNAs, hairpin RNAs, and artificial microRNAs], nonviral genes (nucleases, antiviral inhibitors, and plantibodies), and host-derived resistance genes (dominant resistance genes and recessive resistance genes), and various factors involved in host defense responses. This review examines the above range of approaches used, the viruses that were tested, and the host species that have been examined for resistance, in many cases describing differences in results that were obtained for various systems developed in the last 20 years. We hope this compilation of experiences will aid those who are seeking to use this technology to provide resistance in yet other crops, where nature has not provided such.

  16. Transgenic resistance.

    PubMed

    Cillo, Fabrizio; Palukaitis, Peter

    2014-01-01

    Transgenic resistance to plant viruses is an important technology for control of plant virus infection, which has been demonstrated for many model systems, as well as for the most important plant viruses, in terms of the costs of crop losses to disease, and also for many other plant viruses infecting various fruits and vegetables. Different approaches have been used over the last 28 years to confer resistance, to ascertain whether particular genes or RNAs are more efficient at generating resistance, and to take advantage of advances in the biology of RNA interference to generate more efficient and environmentally safer, novel "resistance genes." The approaches used have been based on expression of various viral proteins (mostly capsid protein but also replicase proteins, movement proteins, and to a much lesser extent, other viral proteins), RNAs [sense RNAs (translatable or not), antisense RNAs, satellite RNAs, defective-interfering RNAs, hairpin RNAs, and artificial microRNAs], nonviral genes (nucleases, antiviral inhibitors, and plantibodies), and host-derived resistance genes (dominant resistance genes and recessive resistance genes), and various factors involved in host defense responses. This review examines the above range of approaches used, the viruses that were tested, and the host species that have been examined for resistance, in many cases describing differences in results that were obtained for various systems developed in the last 20 years. We hope this compilation of experiences will aid those who are seeking to use this technology to provide resistance in yet other crops, where nature has not provided such. PMID:25410101

  17. A comparative study of element concentrations and binding in transgenic and non-transgenic soybean seeds.

    PubMed

    Mataveli, Lidiane Raquel Verola; Pohl, Pawel; Mounicou, Sandra; Arruda, Marco Aurélio Zezzi; Szpunar, Joanna

    2010-12-01

    Transgenic and non-transgenic soybean seeds were compared in terms of total element concentrations, behavior of elements during sequential extraction fractionation and element bioaccessibility using an in vitro simulated gastrointestinal digestion. The analysis were carried out by ICP-sector field-MS or size-exclusion ICP-MS (25 elements in concentrations varying from ng g⁻¹ to the % level). It seems that transgenic and non-transgenic soybean seeds exhibit statistically significant differences in concentrations of Cu, Fe and Sr, which are also reflected by element contents in water extracts and residues. Additionally, contributions of bioaccessible fractions of Cu, Fe and other elements (Mn, S, Zn) for transgenic soybean seeds appear to be larger than those found in non-transgenic soybean seeds.

  18. Insect resistance to Nilaparvata lugens and Cnaphalocrocis medinalis in transgenic indica rice and the inheritance of gna+sbti transgenes.

    PubMed

    Li, Guiying; Xu, Xinping; Xing, Hengtai; Zhu, Huachen; Fan, Qin

    2005-04-01

    Molecular genetic analysis and insect bioassay of transgenic indica rice 'Zhuxian B' plants carrying snowdrop lectin gene (gna) and soybean trypsin inhibitor gene (sbti) were investigated in detail. PCR, 'dot' blot and PCR-Southern blot analysis showed that both transgenes had been incorporated into the rice genome and transmitted up to R3 progeny in most lines tested. Some transgenic lines exhibited Mendelian segregation, but the other showed either 1:1 (positive: negative for the transgenes) or other aberrant segregation patterns. The segregation patterns of gna gene crossed between R2 and R3 progeny. In half of transgenic R3 lines, gna and sbti transgenes co-segregated. Two independent homozygous lines expressing double transgenes were identified in R3 progeny. Southern blot analysis demonstrated that the copy numbers of integrated gna and sbti transgenes varied from one to ten in different lines. Insect bioassay data showed that most transgenic plants had better resistance to both Nilaparvata lugens (Stahl) and Cnaphalocrocis medinalis (Guenee) than wild-type plants. The insect resistance of transgenic lines increased with the increase in transgene positive ratio in most of the transgenic lines. In all, we obtained nine lines of R3 transgenic plants, including one pure line, which had better resistance to both N lugens and C medinalis than wild-type plants.

  19. 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...

  20. 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.

  1. Transgenic models for the study of lung biology and disease.

    PubMed

    Ho, Y S

    1994-04-01

    Transgenic models provide a means of understanding the molecular mechanisms for the temporal, spatial, and stimulus-responsive regulation of gene expression in vivo and importantly the pathophysiological consequences of the altered expression of a normal or mutated gene. To facilitate the application of transgenic models in lung research, this review describes several practical considerations in generation of transgenic mice. The potential of transgenic models in lung research is also illustrated by depicting the current models in lung research including those for understanding lung gene regulation, tumorigenesis, mutation detection, antioxidant defense, emphysema, fibrosis, and hypertension. The impact of important new development of producing transgenic mice carrying large fragments of DNA contained in yeast artificial chromosomes to achieve proper control of transgene expression and gene targeting technology is also discussed. It is anticipated that transgenic models will provide invaluable information in future lung research.

  2. Molecular Analyses of Transgenic Plants.

    PubMed

    Trijatmiko, Kurniawan Rudi; Arines, Felichi Mae; Oliva, Norman; Slamet-Loedin, Inez Hortense; Kohli, Ajay

    2016-01-01

    One of the major challenges in plant molecular biology is to generate transgenic plants that express transgenes stably over generations. Here, we describe some routine methods to study transgene locus structure and to analyze transgene expression in plants: Southern hybridization using DIG chemiluminescent technology for characterization of transgenic locus, SYBR Green-based real-time RT-PCR to measure transgene transcript level, and protein immunoblot analysis to evaluate accumulation and stability of transgenic protein product in the target tissue. PMID:26614292

  3. Studies of an expanded trinucleotide repeat in transgenic mice

    SciTech Connect

    Bingham, P.; Wang, S.; Merry, D.

    1994-09-01

    Spinal and bulbar muscular atrophy (SBMA) is a progressive motor neuron disease caused by expansion of a trinucleotide repeat in the androgen receptor gene (AR{sup exp}). AR{sup exp} repeats expand further or contract in approximately 25% of transmissions. Analogous {open_quotes}dynamic mutations{close_quotes} have been reported in other expanded trinucleotide repeat disorders. We have been developing a mouse model of this disease using a transgenic approach. Expression of the SBMA AR was documented in transgenic mice with an inducible promoter. No phenotypic effects of transgene expression were observed. We have extended our previous results on stability of the expanded trinucleotide repeat in transgenic mice in two lines carrying AR{sup exp}. Tail DNA was amplified by PCR using primers spanning the repeat on 60 AR{sup exp} transgenic mice from four different transgenic lines. Migration of the PCR product through an acrylamide gel showed no change of the 45 CAG repeat length in any progeny. Similarly, PCR products from 23 normal repeat transgenics showed no change from the repeat length of the original construct. Unlike the disease allele in humans, the expanded repeat AR cDNA in transgenic mice showed no change in repeat length with transmission. The relative stability of CAG repeats seen in the transgenic mice may indicate either differences in the fidelity of replicative enzymes, or differences in error identification and repair between mice and humans. Integration site or structural properties of the transgene itself might also play a role.

  4. 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…

  5. Pharming and transgenic plants.

    PubMed

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

    2007-01-01

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

  6. Pharming and transgenic plants.

    PubMed

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

    2007-01-01

    Plant represented the essence of pharmacopoeia until the beginning of the 19th century when plant-derived pharmaceuticals were partly supplanted by drugs produced by the industrial methods of chemical synthesis. In the last decades, genetic engineering has offered an alternative to chemical synthesis, using bacteria, yeasts and animal cells as factories for the production of therapeutic proteins. More recently, molecular farming has rapidly pushed towards plants among the major players in recombinant protein production systems. Indeed, therapeutic protein production is safe and extremely cost-effective in plants. Unlike microbial fermentation, plants are capable of carrying out post-translational modifications and, unlike production systems based on mammalian cell cultures, plants are devoid of human infective viruses and prions. Furthermore, a large panel of strategies and new plant expression systems are currently developed to improve the plant-made pharmaceutical's yields and quality. Recent advances in the control of post-translational maturations in transgenic plants will allow them, in the near future, to perform human-like maturations on recombinant proteins and, hence, make plant expression systems suitable alternatives to animal cell factories. PMID:17875476

  7. An analytical model assessing the potential threat to natural habitats from insect resistance transgenes: continuous transgene input.

    PubMed

    Kelly, Colleen K; Bowler, Michael; Breden, Felix

    2006-06-22

    The potential effects of 'escape' of genetically modified material (transgenes) into natural communities is a major concern in their use. These effects may be limited in the first instance by limiting the proportion of transgene-carrying plants in the natural community. We previously presented an analytical model of the ecological processes governing the relative abundance and persistence of insect resistance (IR) transgenes in a natural community. In that paper, we illustrated the case in which the transgene is input into the community in a single season using data from oilseed rape (OSR) and its known herbivore, Plutella macropennis. We found that the transgene is unlikely to have a great impact on the natural community. Here, we extend the model for repeated input of crop pollen carrying the transgene. We show the model output, again using OSR, for continuous input of the transgene over 10 years, the projected commercial lifetime of a transgene without associated undesirable agronomic effects. Our results do not change our original conclusion that the IR transgene need not have a large impact on the natural community and our suggestions for assessing and mitigating any threat still stand.

  8. 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.

  9. Growth and endocrine effect of growth hormone transgene dosage in diploid and triploid coho salmon.

    PubMed

    Devlin, Robert H; Sakhrani, Dionne; Biagi, Carlo A; Smith, Jack L; Fujimoto, Takafumi; Beckman, Brian

    2014-01-15

    Growth-hormone transgene dosage, polyploidy, and parental effects on growth and endocrine responses have been assessed in coho salmon. Diploid fry with one or two transgene doses grew equally, whereas later-stage juvenile homozygotes grew faster than hemizygotes. In contrast, homozygotes and hemizygotes grew equally after smoltification, both in sea water and fresh water. Triploid transgenic salmon showed impaired growth which could not be fully overcome with additional transgene copies. Levels of muscle GH mRNA were elevated in two vs. one transgene dose diploids, but in triploids, a dosage effect was observed in muscle but not for animals carrying three transgene doses. IGF-I mRNA levels were elevated in transgenic vs. non-transgenic animals, but a dosage effect was not observed. Diploids and triploids with two transgenes had higher plasma GH levels than one-dose animals, but three-dose triploids showed no further elevation. Circulating IGF-I levels also showed a dosage effect in diploids, but not among any transgene doses in triploids. The present study reveals complex interactions among transgene dosage, maternal effects, developmental stage, and ploidy on growth and endocrine parameters in GH transgenic coho salmon. Specifically, GH transgenes do not always express nor have effects on growth that are directly correlated with the number of transgenes. Further, the reduced growth rate seen in triploid transgenic animals could not be fully overcome by increasing transgene dosage. The findings have relevance for understanding growth physiology, transgene function, and for environmental risk assessments that require understanding phenotypes of hemizygous vs. homozygous transgenic animals in populations.

  10. Transgenic Spartina alterniflora for phytoremediation.

    PubMed

    Czakó, Mihály; Feng, Xianzhong; He, Yuke; Liang, Dali; Márton, László

    2006-01-01

    Perennial monoculture forming grasses are very important natural remediators of pollutants. Their genetic improvement is an important task because introduction of key transgenes can dramatically improve their remediation potential. Transfer of key genes for mercury phytoremediation into the salt marsh cordgrass (Spartina alterniflora) is reported here. S. alterniflora plays an important role in the salt marsh by cycling of elements, both nutrients and pollutants, protects the coastline from erosion, is a keystone species in the salt marsh supporting a large food web, which in turn supports a significant segment of economy, including tourism, has an impact on cloud formation and consequently on global weather, and is thus an ecologically important species relevant for our life-support systems. Embryogenic callus of S. alterniflora was co-inoculated with a pair of Agrobacterium strains LBA4404 carrying the organomercurial lyase (merB) and mercuric reductase (merA) genes, respectively, in order to co-introduce both the merA and the merB genes. Seven stable geneticin resistant lines were recovered. The presence of merA and merB genes was verified by PCR and Southern blotting. All but one transgenic lines contained both the merA and the merB sequences proving that co-introduction into Spartina of two genes from separate Agrobacterium strains is feasible and frequent, although the overall frequency of transformation is low. Northern blotting showed differences in relative expression of the two transgenes among individual transformants. The steady-state RNA levels appeared to correlate with the phenotype. Line #7 showed the highest resistance to HgCl(2) (up to 500 microM), whereas line #3 was the most resistant to phenylmercuric acetate (PMA). Wild-type (WT) callus is sensitive to PMA at 50 microM and to HgCl(2) at 225 microM.

  11. Novel mobilizable prokaryotic two-hybrid system vectors for high-throughput protein interaction mapping in Escherichia coli by bacterial conjugation

    PubMed Central

    Clarke, Paul; Cuív, Páraic Ó; O'Connell, Michael

    2005-01-01

    Since its initial description, the yeast two-hybrid (Y2H) system has been widely used for the detection and analysis of protein–protein interactions. Mating-based strategies have been developed permitting its application for automated proteomic interaction mapping projects using both exhaustive and high-throughput strategies. More recently, a number of prokaryotic two-hybrid (P2H) systems have been developed but, despite the many advantages such Escherichia coli-based systems have over the Y2H system, they have not yet been widely implemented for proteomic interaction mapping. This may be largely due to the fact that high-throughput strategies employing bacterial transformation are not as amenable to automation as Y2H mating-based strategies. Here, we describe the construction of novel conjugative P2H system vectors. These vectors carry a mobilization element of the IncPα group plasmid RP4 and can therefore be mobilized with high efficiency from an E.coli donor strain encoding all of the required transport functions in trans. We demonstrate how these vectors permit the exploitation of bacterial conjugation for technically simplified and automated proteomic interaction mapping strategies in E.coli, analogous to the mating-based strategies developed for the Y2H system. PMID:15687376

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

    PubMed

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

    2016-09-01

    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:27381397

  13. "Christian carrying goomies".

    PubMed

    1994-01-01

    Dr. Passingan Usurup tells critics of his pragmatic approach on condom promotion that he is a Christian carrying condoms for Christ. He is head of the University of Papua New Guinea Medical Center and is credited with developing an AIDS/HIV policy for the Papua New Guinea Defence Force. The condoms were named Goomy and promoted at launching in 1992 in a blue packet under the slogan "The bond that guards." Goomy was chosen as the name because it is pidgin for rubber, chewing gum, and anything associated with rubber. Blue packets were chosen over the calls of most soldiers for a camouflage design because of its universal appeal as the color of the sea and sky and because it was the preference of women in the airlines. Once firmly ensconced in his role at the University, Usurup plans to develop a policy for students and staff and help to conduct AIDS prevention and education activities on campus. He will encourage students to test for HIV rather than highlighting the gloom and doom of infection and disease.

  14. 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...

  15. 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

  16. Design and Management of Field Trials of Transgenic Cereals

    NASA Astrophysics Data System (ADS)

    Bedő, Zoltán; Rakszegi, Mariann; Láng, László

    The development of gene transformation systems has allowed the introgression of alien genes into plant genomes, thus providing a mechanism for broadening the genetic resources available to plant breeders. The design and the management of field trials vary according to the purpose for which transgenic cereals are developed. Breeders study the phenotypic and genotypic stability of transgenic plants, monitor the increase in homozygosity of transgenic genotypes under field conditions, and develop backcross generations to transfer the introduced genes into secondary transgenic cereal genotypes. For practical purposes, they may also multiply seed of the transgenic lines to produce sufficient amounts of grain for the detailed analysis of trait(s) of interest, to determine the field performance of transgenic lines, and to compare them with the non-transformed parental genotypes. Prior to variety registration, the Distinctness, Uniformity and Stability (DUS) tests and Value for Cultivation and Use (VCU) experiments are carried out in field trials. Field testing includes specific requirements for transgenic cereals to assess potential environmental risks. The capacity of the pollen to survive, establish and disseminate in the field test environment, the potential for gene transfer, the effects of products expressed by the introduced sequences and phenotypic and genotypic instability that might cause deleterious effects must all be specifically monitored, as required by EU Directives 2003/701/EC (1) on the release of genetically modified higher plants in the environment.

  17. 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.

  18. 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.

  19. Transgenic algae engineered for higher performance

    SciTech Connect

    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.

  20. 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

  1. Ethical issues in transgenics.

    PubMed

    Sherlock, R; Morrey, J D

    2000-01-01

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

  2. 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.

  3. Carrying Position Influences Infant Behavior.

    ERIC Educational Resources Information Center

    Field, Tiffany; And Others

    1996-01-01

    A total of 32 3-month-old infants were carried by their mothers in a soft infant carrier designed to place the infants facing either inward or outward. A within-subject comparison found that when infants were carried facing in, they spent significantly more time sleeping, while infants carried facing out were more active. (MDM)

  4. Monitoring the escape of transgenic oilseed rape around Japanese ports and roadsides.

    PubMed

    Saji, Hikaru; Nakajima, Nobuyoshi; Aono, Mitsuko; Tamaoki, Masanori; Kubo, Akihiro; Wakiyama, Seiji; Hatase, Yoriko; Nagatsu, Masato

    2005-01-01

    An investigation was carried out to monitor the escape and spread of oilseed rape (Brassica napus) transgenic plants and the introgression of transgenes to its closely related feral species in Japan. We screened a total of about 7500 feral B. napus, 300 B. rapa, and 5800 B. juncea seedlings from maternal plants in 143 locations at several ports, roadsides, and riverbanks. The presence of glufosinate-resistance or glyphosate-resistance transgenes in these seedlings was confirmed by means of herbicide treatments and also immunochemical and DNA analyses. B. napus plants with herbicide-resistant transgenic seeds were found at five of six major ports and along two of four sampled roadsides in the Kanto District. Transgenic oilseed rape plants have not been commercially cultivated in Japan, suggesting that the transgenes would probably have come from imported transgenic seeds that were spilled during transportation to oilseed processing facilities. No transgenes were detected in seeds collected from B. napus plants growing along riverbanks in the Kanto District or in seeds from closely related species (B. rapa and B. juncea). To our knowledge, this is the first published example of feral, transgenic populations occurring in a nation where the transgenic crop has not been cultivated commercially.

  5. APP transgenic mice: their use and limitations.

    PubMed

    Balducci, Claudia; Forloni, Gianluigi

    2011-06-01

    Alzheimer's disease is the most widespread form of dementia. Its histopathological hallmarks include vascular and extracellular β-amyloid (Aβ) deposition and intraneuronal neurofibrillary tangles (NFTs). Gradual decline of cognitive functions linked to progressive synaptic loss makes patients unable to store new information in the earlier stages of the pathology, later becoming completely dependent because they are unable to do even elementary daily life actions. Although more than a hundred years have passed since Alois Alzheimer described the first case of AD, and despite many years of intense research, there are still many crucial points to be discovered in the neuropathological pathway. The development of transgenic mouse models engineered with overexpression of the amyloid precursor protein carrying familial AD mutations has been extremely useful. Transgenic mice present the hallmarks of the pathology, and histological and behavioural examination supports the amyloid hypothesis. As in human AD, extracellular Aβ deposits surrounded by activated astrocytes and microglia are typical features, together with synaptic and cognitive defects. Although animal models have been widely used, they are still being continuously developed in order to recapitulate some missing aspects of the disease. For instance, AD therapeutic agents tested in transgenic mice gave encouraging results which, however, were very disappointing in clinical trials. Neuronal cell death and NFTs typical of AD are much harder to replicate in these mice, which thus offer a fundamental but still imperfect tool for understanding and solving dementia pathology.

  6. Transgenic oil palm: production and projection.

    PubMed

    Parveez, G K; Masri, M M; Zainal, A; Majid, N A; Yunus, A M; Fadilah, H H; Rasid, O; Cheah, S C

    2000-12-01

    Oil palm is an important economic crop for Malaysia. Genetic engineering could be applied to produce transgenic oil palms with high value-added fatty acids and novel products to ensure the sustainability of the palm oil industry. Establishment of a reliable transformation and regeneration system is essential for genetic engineering. Biolistic was initially chosen as the method for oil palm transformation as it has been the most successful method for monocotyledons to date. Optimization of physical and biological parameters, including testing of promoters and selective agents, was carried out as a prerequisite for stable transformation. This has resulted in the successful transfer of reporter genes into oil palm and the regeneration of transgenic oil palm, thus making it possible to improve the oil palm through genetic engineering. Besides application of the Biolistics method, studies on transformation mediated by Agrobacterium and utilization of the green fluorescent protein gene as a selectable marker gene have been initiated. Upon the development of a reliable transformation system, a number of useful targets are being projected for oil palm improvement. Among these targets are high-oleate and high-stearate oils, and the production of industrial feedstock such as biodegradable plastics. The efforts in oil palm genetic engineering are thus not targeted as commodity palm oil. Due to the long life cycle of the palm and the time taken to regenerate plants in tissue culture, it is envisaged that commercial planting of transgenic palms will not occur any earlier than the year 2020.

  7. 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...

  8. Stability of transgene methylation patterns in mice: position effects, strain specificity and cellular mosaicism.

    PubMed

    Koetsier, P A; Mangel, L; Schmitz, B; Doerfler, W

    1996-07-01

    The methylation status of a transgene, which carried the adenovirus type 2 E2A late promoter linked to the chloramphenicol acetyltransferase gene, was studied in three transgenic mouse lines (5-8, 7-1 and 8-1). These lines were analysed over a large number of offspring generations beyond the founder animal. In mating experiments, the influence of the parent-of-origin and strain-specific backgrounds on the transgene methylation patterns were assessed and found to have no effect on the pre-established methylation patterns in mouse lines 5-8 and 8-1. The founder animal 7-1 carried two groups of a total of ten transgenes, which were located on two different chromosomes. These arrays of transgenes could be segregated into separate mouse lines 7-1A and 7-1B. The transgenes of 7-1A animals exhibited cellular mosaic methylation patterns that were demethylated in approximately 10% of the offspring in a mixed genetic background. Upon further transmission of these transgenes in a mixed genetic background, the grandparental methylation patterns were reestablished in most progeny. Mating to inbred DBA/2 mice resulted in maintenance of the demethylated pattern or in further demethylation of the transgenes in approximately 50% of the offspring. In contrast, an equal number of transgenic siblings from matings to C57BL/6 mice showed a return to the original methylation pattern. The mosaic methylation status of this locus was apparently controlled by mouse-strain-specific factors. The methylation patterns of the 7-1B transgenes were not cellular mosaic and remained stable in all offspring, as with lines 5-8 and 8-1. Hence, the strain-dependent and cellular mosaic transgene methylation patterns of 7-1A animals were probably a consequence of the chromosomal integration site of the transgenes (position effect).

  9. [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.

  10. 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.

  11. A globin enhancer acts by increasing the proportion of erythrocytes expressing a linked transgene.

    PubMed Central

    Sutherland, H G; Martin, D I; Whitelaw, E

    1997-01-01

    Enhancer elements have been shown to affect the probability of a gene establishing an active transcriptional state and suppress the silencing of reporter genes in cell lines, but their effect in transgenic mice has been obscured by the use of assays that do not assess expression on a cell-by-cell basis. We have examined the effect of a globin enhancer on the variegation of lacZ expression in erythrocytes of transgenic mice. Mice carrying lacZ driven by the alpha-globin promoter exhibit beta-galactosidase (beta-Gal) expression in only a very small proportion of embryonic erythrocytes. When the transgenic construct also contains the (alphaHS-40 enhancer, which controls expression of the alpha-globin gene, expression is seen in a high proportion of embryonic erythrocytes, although there are variations between transgenic lines which can be attributed to different sites of integration. Analysis of beta-Gal expression levels suggests that expressing cells in lines carrying only the alpha-globin promoter express as much beta-Gal as those in which the transgene also contains alphaHS-40. A marked decline in transgene expression occurs as mice age, which is mainly due to a decrease in the proportion of cells expressing the transgene. Thus, a globin enhancer can act to suppress variegation of a linked transgene; this result is consistent with a model in which enhancers act to establish and maintain an active domain without directly affecting the transcriptional rate. PMID:9032288

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

    PubMed Central

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

    2015-01-01

    ABSTRACT 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. PMID:25984767

  13. 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

  14. 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

  15. 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.

  16. Recurrent Selection for Transgene Activity Levels in Maize Results in Proxy Selection for a Native Gene with the Same Promoter.

    PubMed

    Bodnar, Anastasia L; Schroder, Megan N; Scott, M Paul

    2016-01-01

    High activity levels of a transgene can be very useful, making a transgene easier to evaluate for safety and efficacy. High activity levels can also increase the economic benefit of the production of high value proteins in transgenic plants. The goal of this research is to determine if recurrent selection for activity of a transgene will result in higher activity, and if selection for activity of a transgene controlled by a native promoter will also increase protein levels of the native gene with the same promoter. To accomplish this goal we used transgenic maize containing a construct encoding green fluorescent protein controlled by the promoter for the maize endosperm-specific 27 kDa gamma zein seed storage protein. We carried out recurrent selection for fluorescence intensity in two breeding populations. After three generations of selection, both selected populations were significantly more fluorescent and had significantly higher levels of 27 kDa gamma zein than the unselected control populations. These higher levels of the 27 kDa gamma zein occurred independently of the presence of the transgene. The results show that recurrent selection can be used to increase activity of a transgene and that selection for a transgene controlled by a native promoter can increase protein levels of the native gene with the same promoter via proxy selection. Moreover, the increase in native gene protein level is maintained in the absence of the transgene, demonstrating that proxy selection can be used to produce non-transgenic plants with desired changes in gene expression.

  17. High-efficiency transformation by biolistics of soybean, common bean and cotton transgenic plants.

    PubMed

    Rech, Elibio L; Vianna, Giovanni R; Aragão, Francisco J L

    2008-01-01

    This protocol describes a method for high-frequency recovery of transgenic soybean, bean and cotton plants, by combining resistance to the herbicide imazapyr as a selectable marker, multiple shoot induction from embryonic axes of mature seeds and biolistics techniques. This protocol involves the following stages: plasmid design, preparation of soybean, common bean and cotton apical meristems for bombardment, microparticle-coated DNA bombardment of apical meristems and in vitro culture and selection of transgenic plants. The average frequencies (the total number of fertile transgenic plants divided by the total number of bombarded embryonic axes) of producing germline transgenic soybean and bean and cotton plants using this protocol are 9, 2.7 and 0.55%, respectively. This protocol is suitable for studies of gene function as well as the production of transgenic cultivars carrying different traits for breeding programs. This protocol can be completed in 7-10 months.

  18. 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.

  19. A transgenic apple callus showing reduced polyphenol oxidase activity and lower browning potential.

    PubMed

    Murata, M; Nishimura, M; Murai, N; Haruta, M; Homma, S; Itoh, Y

    2001-02-01

    Polyphenol oxidase (PPO) is responsible for enzymatic browning of apples. Apples lacking PPO activity might be useful not only for the food industry but also for studies of the metabolism of polyphenols and the function of PPO. Transgenic apple calli were prepared by using Agrobacterium tumefaciens carrying the kanamycin (KM) resistant gene and antisense PPO gene. Four KM-resistant callus lines were obtained from 356 leaf explants. Among these transgenic calli, three calli grew on the medium containing KM at the same rate as non-transgenic callus on the medium without KM. One callus line had an antisense PPO gene, in which the amount and activity of PPO were reduced to half the amount and activity in non-transgenic callus. The browning potential of this line, which was estimated by adding chlorogenic acid, was also half the browning potential of non-transgenic callus.

  20. 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.

  1. Generation of transgenic energy cane plants with integration of minimal transgene expression cassette.

    PubMed

    Fouad, Walid M; Hao, Wu; Xiong, Yuan; Steeves, Cody; Sandhu, Surinder K; Altpeter, Fredy

    2015-01-01

    Lignocellulosic biomass has the potential to serve as feedstock and direct replacement for petrochemicals in the fuel, chemical, pharmaceutical and material industries. Energy cane has been identified by the U.S. Department of Energy (DOE) as prime lignocellulosic feedstock as it produces record biomass yields and is able to grow on low-value land with reduced inputs. Molecular improvement of energy cane is an essential step toward the development of a high-value crop and may contribute to improved biomass conversion to value added products. Such improvements require a development of an efficient regeneration and transformation system for the vegetatively propagated energy cane varieties. In this report, an efficient biolistic gene delivery protocol for energy canes (genotype L 79-1002 and Ho 00-961) has been established with immature leaf rolls as explants. Embryonic calli, developed approximately 6 weeks after culture initiation and was used as target for biolistic transfer of a minimum expression cassette of P-ubi::nptII::35S polyA derived from plasmid pJFNPTII. Putative transgenic clones of callus were obtained after selection on callus induction medium supplemented with 30 mg l(-1) geneticin. Regeneration was carried out on NB medium, which is modified from MS supplemented with 1.86 mg l(-1) naphthaleneacetic acid (NAA) and 0.1mg l(-1), 6- benzylaminopurine (BAP) and 20mg l(-1) paromomycin. Shoots growing on selection media were transferred to hormone free medium with 20 mg l(-1) paromomycin. Putative transgenic lines were first analyzed by PCR. Transgene integration was confirmed by Southern blot analysis. ELISA (Enzyme-Linked Immunosorbent Assay) and Immunochromathography assays confirmed transgene expression.

  2. 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...

  3. Can transgenic mosquitoes afford the fitness cost?

    PubMed

    Lambrechts, Louis; Koella, Jacob C; Boëte, Christophe

    2008-01-01

    In a recent study, SM1-transgenic Anopheles stephensi, which are resistant partially to Plasmodium berghei, had higher fitness than non-transgenic mosquitoes when they were maintained on Plasmodium-infected blood. This result should be interpreted cautiously with respect to malaria control using transgenic mosquitoes because, despite the evolutionary advantage conferred by the transgene, a concomitant cost prevents it from invading the entire population. Indeed, for the spread of a resistance transgene in a natural situation, the transgene's fitness cost and the efficacy of the gene drive will be more crucial than any evolutionary advantage.

  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. Cryopreservation of transgenic mouse lines.

    PubMed

    Pomeroy, K O

    1993-01-01

    A transgenic animal represents an enormous investment in time and money. Animals can be destroyed through disease, fire, malfuncnons in the control of the environment, negligence, sabotage, or accidental disposal. Researchers can protect valuable transgenic lines from accrdental destruction by "banking" them in liquid nitrogen. Cryopreservation can also reduce animal costs by decreasing the number of live animals investigators must maintain. Often, when one is trying to produce a transgenic animal, some lines will be derived that may not initially appear interesting. These animals can be stored in liquid nitrogen for future recovery and study. The maintenance of just one line of mice, say 25 mice at 15 cents/d, can cost over $1000 (US) in a single year. PMID:21390665

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

    PubMed

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

    2001-02-01

    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.

  7. [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.

  8. Biotransformation of hyoscyamine into scopolamine in transgenic tobacco cell cultures.

    PubMed

    Moyano, Elisabeth; Palazón, Javier; Bonfill, Mercedes; Osuna, Lidia; Cusidó, Rosa M; Oksman-Caldentey, Kirsi-Marja; Piñol, M Teresa

    2007-04-01

    Hyoscyamine-6beta-hydroxylase (H6H) catalyses the conversion of hyoscyamine into its epoxide scopolamine, a compound with a higher added value in the pharmaceutical market than hyoscyamine. We report the establishment of tobacco cell cultures carrying the Hyoscyamus muticus h6h gene under the control of the promoter CAMV 35S. The cell cultures were derived from hairy roots obtained via genetically modified Agrobacterium rhizogenes carrying the pRi and pLAL21 plasmids. The cultures were fed with hyoscyamine, and 4 weeks later the amount of scopolamine produced was quantified by HPLC. The transgenic cell suspension cultures showed a considerable capacity for the bioconversion of hyoscyamine into scopolamine, and released it to the culture medium. Although the scale-up from shake-flask to bioreactor culture usually results in reduced productivities, our transgenic cells grown in a 5-L turbine stirred tank reactor in a batch mode significantly increased the scopolamine accumulation.

  9. 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)

  10. Loren Shriver carries Olympic torch

    NASA Technical Reports Server (NTRS)

    1996-01-01

    KSC Shuttle Operations Manager Loren J. Shriver proudly displays the Olympic torch that he carried to the top of Launch Pad 39A as his contribution to the July 7, 1996 KSC Olympic torch relay effort. Nineteen other KSC runners also participated in the relay effort at the Center. The Olympic torch arrived at KSC at 1:40 p.m. and traveled a 20-mile course to the pad and then out to the KSC visitor Center. The Space Shuttle Atlantis is behind Shriver, poised for the STS-79 mission, which will feature the fourth docking of the Shuttle with the Russian Mir space station.

  11. [Current status and industrialization of transgenic tomatoes].

    PubMed

    Wang, Ao-Xue; Chen, Xiu-Ling

    2011-09-01

    In this review, the progress in transgenic tomato research, including disease and insect resistance, herbicide resistance, stress tolerance, long-term storage, quality improvement, and male sterility, were described. The recent researches on producing heterologous proteins using transgenic tomatoes were also reviewed. Furthermore, the industrialization status and problems of transgenic tomatoes were analyzed and the prospects of both research and industrialization in transgenic tomatoes were discussed.

  12. 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...

  13. [Features of development and reproduction of transgenic flax].

    PubMed

    Lemesh, V A; Samatadze, T E; Guzenko, E V; Zhelezniakova, E V; Amosova, A V; Zelenin, A V; Muravenko, O V

    2014-01-01

    Primary transformants carrying a genetic construct with the chimeric gfp-tua6 gene were obtained using biolistic transformation of hypocotyl explants of flax variety Vasilek. Viable modified plants were used as a basis for the production of inbred lines with confirmed inheritance of introduced genetic construct in three generations. The characteristics of phenological growth stages, plant height, number of bolls and meiosis were studied for transgenic plants. A comparison of transformed lines based on reproduction years revealed a significant decrease of seed production in one line. Meiotic analysis of this line at metaphase I and anaphase I stages was conducted. The percentage of cells with impaired meiosis was highest in transgenic plants of the line with the lowest seed production. Thus, the nonspecific incorporation of genetic construct into the flax genome using biolistic transformation impairs meiosis to a different extent and it is the main reason for unequal reproducibility of transgenic flax. The production of stably reproducing transgenic lines requires systematic analysis of meiosis.

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

    PubMed

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

    2011-07-01

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

  15. Detection of a Common and Persistent tet(L)-Carrying Plasmid in Chicken-Waste-Impacted Farm Soil

    PubMed Central

    Hilpert, Markus; Ward, Mandy J.

    2012-01-01

    The connection between farm-generated animal waste and the dissemination of antibiotic resistance in soil microbial communities, via mobile genetic elements, remains obscure. In this study, electromagnetic induction (EMI) surveying of a broiler chicken farm assisted soil sampling from a chicken-waste-impacted site and a marginally affected site. Consistent with the EMI survey, a disparity existed between the two sites with regard to soil pH, tetracycline resistance (Tcr) levels among culturable soil bacteria, and the incidence and prevalence of several tet and erm genes in the soils. No significant difference was observed in these aspects between the marginally affected site and several sites in a relatively pristine regional forest. When the farm was in operation, tet(L), tet(M), tet(O), erm(A), erm(B), and erm(C) genes were detected in the waste-affected soil. Two years after all waste was removed from the farm, tet(L), tet(M), tet(O), and erm(C) genes were still detected. The abundances of tet(L), tet(O), and erm(B) were measured using quantitative PCR, and the copy numbers of each were normalized to eubacterial 16S rRNA gene copy numbers. tet(L) was the most prevalent gene, whereas tet(O) was the most persistent, although all declined over the 2-year period. A mobilizable plasmid carrying tet(L) was identified in seven of 14 Tcr soil isolates. The plasmid's hosts were identified as species of Bhargavaea, Sporosarcina, and Bacillus. The plasmid's mobilization (mob) gene was quantified to estimate its prevalence in the soil, and the ratio of tet(L) to mob was shown to have changed from 34:1 to 1:1 over the 2-year sampling period. PMID:22389375

  16. Strain-specific regulation of striatal phenotype in Drd2-eGFP BAC transgenic mice.

    PubMed

    Chan, C Savio; Peterson, Jayms D; Gertler, Tracy S; Glajch, Kelly E; Quintana, Ruth E; Cui, Qiaoling; Sebel, Luke E; Plotkin, Joshua L; Shen, Weixing; Heiman, Myriam; Heintz, Nathaniel; Greengard, Paul; Surmeier, D James

    2012-07-01

    Mice carrying bacterial artificial chromosome (BAC) transgenes have become important tools for neuroscientists, providing a powerful means of dissecting complex neural circuits in the brain. Recently, it was reported that one popular line of these mice--mice possessing a BAC transgene with a D(2) dopamine receptor (Drd2) promoter construct coupled to an enhanced green fluorescent protein (eGFP) reporter--had abnormal striatal gene expression, physiology, and motor behavior. Unlike most of the work using BAC mice, this interesting study relied upon mice backcrossed on the outbred Swiss Webster (SW) strain that were homozygous for the Drd2-eGFP BAC transgene. The experiments reported here were conducted to determine whether mouse strain or zygosity was a factor in the reported abnormalities. As reported, SW mice were very sensitive to transgene expression. However, in more commonly used inbred strains of mice (C57BL/6, FVB/N) that were hemizygous for the transgene, the Drd2-eGFP BAC transgene did not alter striatal gene expression, physiology, or motor behavior. Thus, the use of inbred strains of mice that are hemizygous for the Drd2 BAC transgene provides a reliable tool for studying basal ganglia function.

  17. Curcumin therapy in a Plp1 transgenic mouse model of Pelizaeus-Merzbacher disease

    PubMed Central

    Epplen, Dirk B; Prukop, Thomas; Nientiedt, Tobias; Albrecht, Philipp; Arlt, Friederike A; Stassart, Ruth M; Kassmann, Celia M; Methner, Axel; Nave, Klaus-Armin; Werner, Hauke B; Sereda, Michael W

    2015-01-01

    Objective Pelizaeus–Merzbacher disease (PMD) is a progressive and lethal leukodystrophy caused by mutations affecting the proteolipid protein (PLP1) gene. The most common cause of PMD is a duplication of PLP1 and at present there is no curative therapy available. Methods By using transgenic mice carrying additional copies of Plp1, we investigated whether curcumin diet ameliorates PMD symptoms. The diet of Plp1 transgenic mice was supplemented with curcumin for 10 consecutive weeks followed by phenotypical, histological and immunohistochemical analyses of the central nervous system. Plp1 transgenic and wild-type mice fed with normal chow served as controls. Results Curcumin improved the motor phenotype performance of Plp1 transgenic mice by 50% toward wild-type level and preserved myelinated axons by 35% when compared to Plp1 transgenic controls. Furthermore, curcumin reduced astrocytosis, microgliosis and lymphocyte infiltration in Plp1 transgenic mice. Curcumin diet did not affect the pathologically increased Plp1 mRNA abundance. However, high glutathione levels indicating an oxidative misbalance in the white matter of Plp1 transgenic mice were restored by curcumin treatment. Interpretation Curcumin may potentially serve as an antioxidant therapy of PMD caused by PLP1 gene duplication. PMID:26339673

  18. 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.

  19. Lactation induction as a predictor of post-parturition transgene expression in bovine milk.

    PubMed

    Powell, Ann; Kerr, David; Guthrie, David; Wall, Robert

    2007-05-01

    The bovine's long generation interval results in a delay of several years when evaluating mammary specific transgenes in genetically engineered animals. This experiment was conducted to evaluate the feasibility of reducing that waiting period. Lactation was induced in prepubertal bull and heifer calves as a means of predicting transgene behaviour during subsequent post-parturient lactations in the heifers themselves, and in daughters sired by the bulls. The animals carry a lactation-specific transgene encoding lysostaphin, an antimicrobial protein that kills Staphlococcus aureus, a mastitis-causing pathogen. Oestrogen, progesterone and dexamethasone were administered as previously described (Ball et al. 2000) to nine heifers (five transgenics) ranging in weight from 80 to 145 kg. Eight bull calves (seven transgenics) weighing 81-178 kg received additional oestrogen and progesterone injection prior to dexamethasone treatment. All nine heifers responded to the milk induction scheme yielding between 19 ml and 4.5 l over 5 d. Milk volume from the four responding males (30 microl to 2.5 ml) was significantly less than that harvested from females (P=0.025). Only bull calves >117 kg had a positive response. Lysostaphin was detected in all transgenic prepubertal heifers and in two transgenic prepubertal bull calves induced. A positive relationship was observed between lysostaphin's stapholytic activity in the two types of lactations (r2=0.907, P<0.001) thus providing a useful means of predicting subsequent lysostaphin production in post-partum milk.

  20. 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.

  1. Rearrangement and expression of endogenous immunoglobulin genes occur in many murine B cells expressing transgenic membrane IgM.

    PubMed

    Stall, A M; Kroese, F G; Gadus, F T; Sieckmann, D G; Herzenberg, L A; Herzenberg, L A

    1988-05-01

    Transgenic mice carrying immunoglobulin genes coding for mu heavy chain and kappa light chain have been used to study the mechanisms involved in allelic and isotypic exclusion. We report here that individual cells from transgenic mice carrying a functionally rearranged mu heavy chain gene (capable of generating both membrane and secreted forms of IgM) can rearrange an endogenous mu heavy chain gene and simultaneously produce both transgenic and endogenous IgM. These "double-producing" cells express both endogenous and transgenic IgM in the cytoplasm (detected by immunohistology) and on the cell surface (detected by multiparameter fluorescence-activated cell sorter analysis). In addition, they secrete mixed IgM molecules containing both transgenic and endogenous mu heavy chains (detected in serum by radioimmune assay). The transgenic mice studied also have relatively large numbers of cells that produce endogenous immunoglobulin in the absence of detectable transgenic immunoglobulin ("endogenous-only cells"). The mechanisms that generate double-producing cells and endogenous-only cells appear to be under genetic control because the frequencies of these B-cell populations are characteristic for a given transgenic line. Thus, our findings indicate that more is involved in triggering allelic exclusion than the simple presence or absence of membrane mu heavy chains (as has been previously postulated).

  2. 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.

  3. 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.

  4. Nas transgenic mouse line allows visualization of Notch pathway activity in vivo

    PubMed Central

    Souilhol, Céline; Cormier, Sarah; Monet, Marie; Vandormael-Pournin, Sandrine; Joutel, Anne; Babinet, Charles; Cohen-Tannoudji, Michel

    2006-01-01

    The Notch signalling pathway plays multiple and important roles in mammals. However, several aspects of its action, in particular the precise mapping of its sites of activity, remain unclear. To address this issue, we have generated a transgenic line carrying a construct consisting of a nls-lacZ reporter gene under the control of a minimal promoter and multiple RBP-Jκ binding sites. Here we show that this transgenic line, we named NAS for Notch Activity Sensor, displays an expression profile that is consistent with current knowledge on Notch activity sites in mice, even though it may not report on all these sites. Moreover, we observe that NAS transgene expression is abolished in a RBP-Jκ deficient background indicating that it indeed requires Notch/RBP-Jκ signalling pathway activity. Thus, the NAS transgenic line constitutes a valuable and versatile tool to gain further insights into the complex and various functions of the Notch signalling pathway. PMID:16708386

  5. Pancreatic expression of human insulin gene in transgenic mice.

    PubMed

    Bucchini, D; Ripoche, M A; Stinnakre, M G; Desbois, P; Lorès, P; Monthioux, E; Absil, J; Lepesant, J A; Pictet, R; Jami, J

    1986-04-01

    We have investigated the possibility of obtaining integration and expression of a native human gene in transgenic mice. An 11-kilobase (kb) human chromosomal DNA fragment including the insulin gene (1430 base pairs) was microinjected into fertilized mouse eggs. This fragment was present in the genomic DNA of several developing animals. One transgenic mouse and its progeny were analyzed for expression of the foreign gene. Synthesis and release of human insulin was revealed by detection of the human C-peptide in the plasma and urine. Human insulin mRNA was found in pancreas but not in other tissues. These findings indicate that the 11-kb human DNA fragment carries the sequences necessary for tissue-specific expression of the insulin gene and the human regulatory sequences react to homologous signals in the mouse.

  6. Polyhydroxybutyrate synthesis in transgenic flax.

    PubMed

    Wróbel, Magdalena; Zebrowski, Jacek; Szopa, Jan

    2004-01-01

    Flax (Linum usitatissimum L.) is an annual plant species widely cultivated in temperate climates for bast fibres and linseed oil. Apart from traditional textile use, the fibres are fast becoming an integral part of new composite materials utilized in automobile and constructive industry. Especially attractive for environmental safety demands are biodegradable and renewable biocomposities based on polyhydroxybutyrate (PHB) polymer as a matrix and reinforced with the flax fibres. Manufacturing of PHB by bacteria fermentation is however substantially more expansive as compared to technologies producing conventional plastics. We report for the first time generation of transgenic plants which produce both components of flax/PHB composites, i.e. the fibres and the thermoplastic matrix in the same plant organ of a crop. The flax (cv. Nike) plants were transformed using constructs bearing either single cDNA, encoding the beta-ketothiolase enzyme (C plants), or all three of the genes necessary for poly-beta-hydroxybutyrate (PHB) synthesis (M plants). Both constructs contained a plastidial targeting sequence. The amount of PHB produced by the transgenic plants was up to over 70-fold higher than in wild-type plants, when analysed using the gas chromatography/mass spectrometry (GC-MS method). The PHB accumulation in plastids caused change both in their shape and size. The use of a stem-specific promoter for transgene expression protected the transgenic plant from growth retardation and also provided higher PHB synthesis than in the case of constructs governed by the 35S CaMV constitutive promoter. None toxic effects that could lead to stunted growth or the loss of fertility were observed, when 14-3-3 promoter was used as the stem-specific. Significant modifications in stem mechanical properties were accompanied to the PHB accumulation in growing cell of fibres in the transgenic plants. The Young's modulus E, the average measure of stem tissues resistance to tensile loads

  7. Dual promoter lentiviral vector generates transgenic mice expressing E2-CSFV glycoprotein in their milk, but impairs early identification of transgenic embryos.

    PubMed

    Ramos, Oliberto Sánchez; Carratalá, Yanet Prieto; Puerta, Silvia Gómez; Pereira, Natalie C Parra; Amarán, Lester Suárez; Chaves, Silvana P Jimenez; Alonso, Jorge R Toledo

    2011-04-15

    Lentiviral vectors containing the green fluorescent protein gene have been successfully used to select transgenic embryos before transfer to a surrogate mother. However, there are apparently no reports regarding early detection of transgenic embryos using a lentiviral vector carrying an additional transcription unit for tissue-specific expression of a valuable protein. In this study, two HIV-based lentiviral vectors were constructed. The first one contained the green fluorescent protein (GFP) coding sequence driven by the early SV40 promoter (Lv-G), whereas the other contained an additional transcription unit for the expression of E2 glycoprotein from classical swine fever virus, driven by a 1.5 kb αS1casein promoter from water buffalo (Lv-αS1cE2hisG). Microinjection of single-cell mouse embryos with Lv-G lentiviral vector rendered embryos which were GFP-positive, beginning at the four-cell stage. Of 33 mice born, 28 (81%) carried the transgene DNA and 15 (55.5%) were GFP-positive. Microinjection of Lv-αS1cE2hisG lentiviral vector yielded 28 mice born; although 24 (85%) carried the transgene DNA, none were GFP-positive, suggesting that the tissue-specific expression cassette interfered with expression of the ubiquitous trancriptional unit. In Lv-αS1cE2hisG transgenic mice, E2his was expressed in milk as a homodimer (at concentrations ≤ 0.422 mg/mL). This was apparently the first report of expression of a recombinant protein in the milk of transgenic animals generated by lentiviral transgenesis.

  8. Transgene integration: use of matrix attachment regions.

    PubMed

    Allen, George C; Spiker, Steven; Thompson, William F

    2005-01-01

    Matrix attachment regions (MARs) are operationally defined as DNA elements that bind specifically to the nuclear matrix in vitro. When MARs are positioned at the 5'- and 3'-ends of a transgene higher more predictable expression of the transgene results. MARs are increasingly being applied to prevent unwanted transgene silencing, which is especially common when direct DNA transformation methods are used. This chapter describes methods for the isolation of MARs and the subsequent methods allowing the investigator to incorporate MARS into transformation strategies that can both improve transformation frequency and result in predictable, stable expression of the transgenic trait.

  9. Transgene integration: use of matrix attachment regions.

    PubMed

    Allen, George C; Spiker, Steven; Thompson, William F

    2005-01-01

    Matrix attachment regions (MARs) are operationally defined as DNA elements that bind specifically to the nuclear matrix in vitro. When MARs are positioned at the 5'- and 3'-ends of a transgene higher more predictable expression of the transgene results. MARs are increasingly being applied to prevent unwanted transgene silencing, which is especially common when direct DNA transformation methods are used. This chapter describes methods for the isolation of MARs and the subsequent methods allowing the investigator to incorporate MARS into transformation strategies that can both improve transformation frequency and result in predictable, stable expression of the transgenic trait. PMID:15310930

  10. A built-in strategy for containment of transgenic plants: creation of selectively terminable transgenic rice.

    PubMed

    Lin, Chaoyang; Fang, Jun; Xu, Xiaoli; Zhao, Te; Cheng, Jiaan; Tu, Juming; Ye, Gongyin; Shen, Zhicheng

    2008-01-01

    Plant transgenic technology has been widely utilized for engineering crops for trait improvements and for production of high value proteins such as pharmaceuticals. However, the unintended spreading of commercial transgenic crops by pollination and seed dispersal is a major concern for environmental and food safety. Simple and reliable containment strategies for transgenes are highly desirable. Here we report a novel method for creating selectively terminable transgenic rice. In this method, the gene(s) of interest is tagged with a RNA interference cassette, which specifically suppresses the expression of the bentazon detoxification enzyme CYP81A6 and thus renders transgenic rice to be sensitive to bentazon, a herbicide used for rice weed control. We generated transgenic rice plants by this method using a new glyphosate resistant 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) gene from Pesudomonas putida as the gene of interest, and demonstrated that these transgenic rice plants were highly sensitive to bentazon but tolerant to glyphosate, which is exactly the opposite of conventional rice. Field trial of these transgenic rice plants further confirmed that they can be selectively killed at 100% by one spray of bentazon at a regular dose used for conventional rice weed control. Furthermore, we found that the terminable transgenic rice created in this study shows no difference in growth, development and yield compared to its non-transgenic control. Therefore, this method of creating transgenic rice constitutes a novel strategy of transgene containment, which appears simple, reliable and inexpensive for implementation. PMID:18350155

  11. A Built-In Strategy for Containment of Transgenic Plants: Creation of Selectively Terminable Transgenic Rice

    PubMed Central

    Zhao, Te; Cheng, Jiaan; Tu, Juming; Ye, Gongyin; Shen, Zhicheng

    2008-01-01

    Plant transgenic technology has been widely utilized for engineering crops for trait improvements and for production of high value proteins such as pharmaceuticals. However, the unintended spreading of commercial transgenic crops by pollination and seed dispersal is a major concern for environmental and food safety. Simple and reliable containment strategies for transgenes are highly desirable. Here we report a novel method for creating selectively terminable transgenic rice. In this method, the gene(s) of interest is tagged with a RNA interference cassette, which specifically suppresses the expression of the bentazon detoxification enzyme CYP81A6 and thus renders transgenic rice to be sensitive to bentazon, a herbicide used for rice weed control. We generated transgenic rice plants by this method using a new glyphosate resistant 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) gene from Pesudomonas putida as the gene of interest, and demonstrated that these transgenic rice plants were highly sensitive to bentazon but tolerant to glyphosate, which is exactly the opposite of conventional rice. Field trial of these transgenic rice plants further confirmed that they can be selectively killed at 100% by one spray of bentazon at a regular dose used for conventional rice weed control. Furthermore, we found that the terminable transgenic rice created in this study shows no difference in growth, development and yield compared to its non-transgenic control. Therefore, this method of creating transgenic rice constitutes a novel strategy of transgene containment, which appears simple, reliable and inexpensive for implementation. PMID:18350155

  12. A built-in strategy for containment of transgenic plants: creation of selectively terminable transgenic rice.

    PubMed

    Lin, Chaoyang; Fang, Jun; Xu, Xiaoli; Zhao, Te; Cheng, Jiaan; Tu, Juming; Ye, Gongyin; Shen, Zhicheng

    2008-01-01

    Plant transgenic technology has been widely utilized for engineering crops for trait improvements and for production of high value proteins such as pharmaceuticals. However, the unintended spreading of commercial transgenic crops by pollination and seed dispersal is a major concern for environmental and food safety. Simple and reliable containment strategies for transgenes are highly desirable. Here we report a novel method for creating selectively terminable transgenic rice. In this method, the gene(s) of interest is tagged with a RNA interference cassette, which specifically suppresses the expression of the bentazon detoxification enzyme CYP81A6 and thus renders transgenic rice to be sensitive to bentazon, a herbicide used for rice weed control. We generated transgenic rice plants by this method using a new glyphosate resistant 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) gene from Pesudomonas putida as the gene of interest, and demonstrated that these transgenic rice plants were highly sensitive to bentazon but tolerant to glyphosate, which is exactly the opposite of conventional rice. Field trial of these transgenic rice plants further confirmed that they can be selectively killed at 100% by one spray of bentazon at a regular dose used for conventional rice weed control. Furthermore, we found that the terminable transgenic rice created in this study shows no difference in growth, development and yield compared to its non-transgenic control. Therefore, this method of creating transgenic rice constitutes a novel strategy of transgene containment, which appears simple, reliable and inexpensive for implementation.

  13. 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.

  14. Recurrent Selection for Transgene Activity Levels in Maize Results in Proxy Selection for a Native Gene with the Same Promoter

    PubMed Central

    Bodnar, Anastasia L.; Schroder, Megan N.; Scott, M. Paul

    2016-01-01

    High activity levels of a transgene can be very useful, making a transgene easier to evaluate for safety and efficacy. High activity levels can also increase the economic benefit of the production of high value proteins in transgenic plants. The goal of this research is to determine if recurrent selection for activity of a transgene will result in higher activity, and if selection for activity of a transgene controlled by a native promoter will also increase protein levels of the native gene with the same promoter. To accomplish this goal we used transgenic maize containing a construct encoding green fluorescent protein controlled by the promoter for the maize endosperm-specific 27kDa gamma zein seed storage protein. We carried out recurrent selection for fluorescence intensity in two breeding populations. After three generations of selection, both selected populations were significantly more fluorescent and had significantly higher levels of 27kDa gamma zein than the unselected control populations. These higher levels of the 27kDa gamma zein occurred independently of the presence of the transgene. The results show that recurrent selection can be used to increase activity of a transgene and that selection for a transgene controlled by a native promoter can increase protein levels of the native gene with the same promoter via proxy selection. Moreover, the increase in native gene protein level is maintained in the absence of the transgene, demonstrating that proxy selection can be used to produce non-transgenic plants with desired changes in gene expression. PMID:26895451

  15. Transgenic Models in Retinoblastoma Research.

    PubMed

    Nair, Rohini M; Vemuganti, Geeta K

    2015-04-01

    Understanding the mechanism of retinoblastoma (Rb) tumor initiation, development, progression and metastasis in vivo mandates the use of animal models that mimic this intraocular tumor in its genetic, anatomic, histologic and ultrastructural features. An early setback for developing mouse Rb models was that Rb mutations did not cause tumorigenesis in murine retinas. Subsequently, the discovery that the p107 protein takes over the role of pRb in mice led to the development of several animal models that phenotypically and histologically resemble the human form. This paper summarizes the transgenic models that have been developed over the last three decades. PMID:27171579

  16. A foreign dihydrofolate reductase gene in transgenic mice acts as a dominant mutation.

    PubMed Central

    Gordon, J W

    1986-01-01

    We have produced 17 lines of transgenic mice by microinjecting a full-length cDNA clone of an altered dihydrofolate reductase (dhfr) gene. The protein specified by this gene carries a point mutation which triples its Km for dihydrofolate and reduces substrate turnover 20-fold relative to the wild-type enzyme. Transgenic mice from different pedigrees, several of which carry a single copy of this gene in different integration sites, manifest an array of similar developmental abnormalities including growth stunting, reduced fertility, pigmentation changes, and skeletal defects. These defects appear in animals heterozygous for the foreign gene. RNA analyses demonstrate significant expression of the cDNA in newborn mice and adult tissues. These findings show that the additional dhfr gene exerts its mutational effects in a dominant fashion, and therefore the data indicate that transgenic mice can serve as models for elucidating mechanisms of dominant mutagenesis. Images PMID:3785192

  17. Production of transgenic livestock: promise fulfilled.

    PubMed

    Wheeler, M B

    2003-01-01

    The introduction of specific genes into the genome of farm animals and its stable incorporation into the germ line has been a major technological advance in agriculture. Transgenic technology provides a method to rapidly introduce "new" genes into cattle, swine, sheep, and goats without crossbreeding. It is a more extreme methodology, but in essence, not really different from crossbreeding or genetic selection in its result. Methods to produce transgenic animals have been available for more than 20 yr, yet recently lines of transgenic livestock have been developed that have the potential to improve animal agriculture and benefit producers and/or consumers. There are a number of methods that can be used to produce transgenic animals. However, the primary method to date has been the microinjection of genes into the pronuclei of zygotes. This method is one of an array of rapidly developing transgenic methodologies. Another method that has enjoyed recent success is that of nuclear transfer or "cloning." The use of this technique to produce transgenic livestock will profoundly affect the use of transgenic technology in livestock production. Cell-based, nuclear transfer or cloning strategies have several distinct advantages for use in the production of transgenic livestock that cannot be attained using pronuclear injection of DNA. Practical applications of transgenesis in livestock production include enhanced prolificacy and reproductive performance, increased feed utilization and growth rate, improved carcass composition, improved milk production and/or composition, and increased disease resistance. One practical application of transgenics in swine production is to improve milk production and/or composition. To address the problem of low milk production, transgenic swine over-expressing the milk protein bovine alpha-lactalbumin were developed and characterized. The outcomes assessed were milk composition, milk yield, and piglet growth. Our results indicate that

  18. 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…

  19. Expression of an endochitinase gene from Trichoderma virens confers enhanced tolerance to Alternaria blight in transgenic Brassica juncea (L.) czern and coss lines.

    PubMed

    Kamble, Suchita; Mukherjee, Prasun K; Eapen, Susan

    2016-01-01

    An endochitinase gene 'ech42' from the biocontrol fungus 'Trichoderma virens' was introduced to Brassica juncea (L). Czern and Coss via Agrobaterium tumefaciens mediated genetic transformation method. Integration and expression of the 'ech42' gene in transgenic lines were confirmed by PCR, RT-PCR and Southern hybridization. Transgenic lines (T1) showed expected 3:1 Mendelian segregation ratio when segregation analysis for inheritance of transgene 'hpt' was carried out. Fluorimetric analysis of transgenic lines (T0 and T1) showed 7 fold higher endochitinase activity than the non-transformed plant. Fluorimetric zymogram showed presence of endochitinase (42 kDa) in crude protein extract of transgenic lines. In detached leaf bioassay with fungi Alternaria brassicae and Alternaria brassicicola, transgenic lines (T0 and T1) showed delayed onset of lesions as well as 30-73 % reduction in infected leaf area compared to non-transformed plant. PMID:27186020

  20. 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.

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

    PubMed Central

    2012-01-01

    Background While the genetic transformation of the major cereal crops has become relatively routine, to date only a few reports were published on transgenic triticale, and robust data on T-DNA integration and segregation have not been available in this species. Results Here, we present a comprehensive analysis of stable transgenic winter triticale cv. Bogo carrying the selectable marker gene HYGROMYCIN PHOSPHOTRANSFERASE (HPT) and a synthetic green fluorescent protein gene (gfp). Progeny of four independent transgenic plants were comprehensively investigated with regard to the number of integrated T-DNA copies, the number of plant genomic integration loci, the integrity and functionality of individual T-DNA copies, as well as the segregation of transgenes in T1 and T2 generations, which also enabled us to identify homozygous transgenic lines. The truncation of some integrated T-DNAs at their left end along with the occurrence of independent segregation of multiple T-DNAs unintendedly resulted in a single-copy segregant that is selectable marker-free and homozygous for the gfp gene. The heritable expression of gfp driven by the maize UBI-1 promoter was demonstrated by confocal laser scanning microscopy. Conclusions The used transformation method is a valuable tool for the genetic engineering of triticale. Here we show that comprehensive molecular analyses are required for the correct interpretation of phenotypic data collected from the transgenic plants. PMID:23006412

  2. 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

  3. Allergenicity assessment of the papaya ringspot virus coat protein expressed in transgenic rainbow papaya.

    PubMed

    Fermín, Gustavo; Keith, Ronald C; Suzuki, Jon Y; Ferreira, Stephen A; Gaskill, Douglas A; Pitz, Karen Y; Manshardt, Richard M; Gonsalves, Dennis; Tripathi, Savarni

    2011-09-28

    The virus-resistant, transgenic commercial papaya Rainbow and SunUp (Carica papaya L.) have been consumed locally in Hawaii and elsewhere in the mainland United States and Canada since their release to planters in Hawaii in 1998. These papaya are derived from transgenic papaya line 55-1 and carry the coat protein (CP) gene of papaya ringspot virus (PRSV). The PRSV CP was evaluated for potential allergenicity, an important component in assessing the safety of food derived from transgenic plants. The transgene PRSV CP sequence of Rainbow papaya did not exhibit greater than 35% amino acid sequence homology to known allergens, nor did it have a stretch of eight amino acids found in known allergens which are known common bioinformatic methods used for assessing similarity to allergen proteins. PRSV CP was also tested for stability in simulated gastric fluid and simulated intestinal fluid and under various heat treatments. The results showed that PRSV CP was degraded under conditions for which allergenic proteins relative to nonallergens are purported to be stable. The potential human intake of transgene-derived PRSV CP was assessed by measuring CP levels in Rainbow and SunUp along with estimating the fruit consumption rates and was compared to potential intake estimates of PRSV CP from naturally infected nontransgenic papaya. Following accepted allergenicity assessment criteria, our results show that the transgene-derived PRSV CP does not pose a risk of food allergy.

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

    PubMed

    Guttikonda, Satish K; Marri, Pradeep; 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

  5. 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

  6. 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

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

    PubMed

    Guttikonda, Satish K; Marri, Pradeep; 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.

  8. 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

  9. Transgenic animals: current and alternative strategies.

    PubMed

    Chan, A W

    1999-01-01

    Transgenic animal technology is one of the most fascinating technologies developed in the last two decades. It allows us to address questions in life sciences that no other methods have achieved. The impact on biomedical and biological research, as well as commercial interests are overwhelming. The questions accompanying this fast growing technology and its diversified applications attract the attention from a variety of entities. Still, one of the most fundamental problems remaining is the search for an efficient and reliable gene delivery system for creating transgenic animals. The traditional method of pronuclear microinjection has displayed great variability in success among species. While an acceptable efficiency in the production of transgenic mice has been attained, the relative low efficiency (<1%) in creating transgenic livestock has become one of the barriers for its application. In the past decades, improvements in producing transgenic livestock have made a slow progression, however, the recent advancement in cloning technology and the ability to create transgenic livestock in a highly efficient manner, have opened the gate to a new era in transgenic technology. Discoveries of new gene delivery systems have created an enthusiastic atmosphere that has made this technology so unique. This review focuses on gene delivery strategies as well as various approaches that may assist the advancement of transgenic efficiency in large animals.

  10. [New advances in animal transgenic technology].

    PubMed

    Sun, Zhen-Hong; Miao, Xiang-Yang; Zhu, Rui-Liang

    2010-06-01

    Animal transgenic technology is one of the fastest growing biotechnology in the 21st century. It is used to integrate foreign genes into the animal genome by genetic engineering technology so that foreign genes can be expressed and inherited to the offspring. The transgenic efficiency and precise control of gene expression are the key limiting factors on preparation of transgenic animals. A variety of transgenic techniques are available, each of which has its own advantages and disadvantages and still needs further study because of unresolved technical and safety issues. With the in-depth research, the transgenic technology will have broad application prospects in the fields of exploration of gene function, animal genetic improvement, bioreactor, animal disease models, organ transplantation and so on. This article reviews the recently developed animal gene transfer techniques, including germline stem cell mediated method to improve the efficiency, gene targeting to improve the accuracy, RNA interference (RNAi)-mediated gene silencing technology, and the induced pluripotent stem cells (iPS) transgenic technology. The new transgenic techniques can provide a better platform for the study of trans-genic animals and promote the development of medical sciences, livestock production, and other fields.

  11. [Progress in transgenic fish techniques and application].

    PubMed

    Ye, Xing; Tian, Yuan-Yuan; Gao, Feng-Ying

    2011-05-01

    Transgenic technique provides a new way for fish breeding. Stable lines of growth hormone gene transfer carps, salmon and tilapia, as well as fluorescence protein gene transfer zebra fish and white cloud mountain minnow have been produced. The fast growth characteristic of GH gene transgenic fish will be of great importance to promote aquaculture production and economic efficiency. This paper summarized the progress in transgenic fish research and ecological assessments. Microinjection is still the most common used method, but often resulted in multi-site and multi-copies integration. Co-injection of transposon or meganuclease will greatly improve the efficiency of gene transfer and integration. "All fish" gene or "auto gene" should be considered to produce transgenic fish in order to eliminate misgiving on food safety and to benefit expression of the transferred gene. Environmental risk is the biggest obstacle for transgenic fish to be commercially applied. Data indicates that transgenic fish have inferior fitness compared with the traditional domestic fish. However, be-cause of the genotype-by-environment effects, it is difficult to extrapolate simple phenotypes to the complex ecological interactions that occur in nature based on the ecological consequences of the transgenic fish determined in the laboratory. It is critical to establish highly naturalized environments for acquiring reliable data that can be used to evaluate the environ-mental risk. Efficacious physical and biological containment strategies remain to be crucial approaches to ensure the safe application of transgenic fish technology.

  12. 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.

  13. Transgene expression in pear (Pyrus communis L.) driven by a phloem-specific promoter

    Technology Transfer Automated Retrieval System (TEKTRAN)

    A gene expression cassette carrying ß-glucuronidase (uidA) reporter gene under the control of the promoter of the Arabidopsis sucrose-H+ symporter gene (AtSUC2) was introduced to pear plants via an Agrobacterium-mediated leaf-explant transformation procedure. Transgenic shoots were regenerated from...

  14. [Production of transgenic sugarbeet plants (Beta vulgaris L.) using Agrobacterium rhizogenes].

    PubMed

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

    2005-01-01

    Normal phenotype sugarbeet plants transformed with Agrobacterium rhizogenes were produced using direct regeneration from explants without hairy root phase. Kanamycin resistant plants and Ri-roots carrying the genes of neomycin phosphotransferase II and b-glucuronidase have been obtained. Integration of transgenes into sugarbeet genome was confirmed with GUS-assay and PCR using primers for the introduced genes. PMID:16018172

  15. 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

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

    PubMed

    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

  17. 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...

  18. Rapid characterization of transgenic and non-transgenic soybean oils by chemometric methods using NIR spectroscopy.

    PubMed

    Luna, Aderval S; da Silva, Arnaldo P; Pinho, Jéssica S A; Ferré, Joan; Boqué, Ricard

    2013-01-01

    Near infrared (NIR) spectroscopy and multivariate classification were applied to discriminate soybean oil samples into non-transgenic and transgenic. Principal Component Analysis (PCA) was applied to extract relevant features from the spectral data and to remove the anomalous samples. The best results were obtained when with Support Vectors Machine-Discriminant Analysis (SVM-DA) and Partial Least Squares-Discriminant Analysis (PLS-DA) after mean centering plus multiplicative scatter correction. For SVM-DA the percentage of successful classification was 100% for the training group and 100% and 90% in validation group for non transgenic and transgenic soybean oil samples respectively. For PLS-DA the percentage of successful classification was 95% and 100% in training group for non transgenic and transgenic soybean oil samples respectively and 100% and 80% in validation group for non transgenic and transgenic respectively. The results demonstrate that NIR spectroscopy can provide a rapid, nondestructive and reliable method to distinguish non-transgenic and transgenic soybean oils.

  19. Rapid characterization of transgenic and non-transgenic soybean oils by chemometric methods using NIR spectroscopy

    NASA Astrophysics Data System (ADS)

    Luna, Aderval S.; da Silva, Arnaldo P.; Pinho, Jéssica S. A.; Ferré, Joan; Boqué, Ricard

    Near infrared (NIR) spectroscopy and multivariate classification were applied to discriminate soybean oil samples into non-transgenic and transgenic. Principal Component Analysis (PCA) was applied to extract relevant features from the spectral data and to remove the anomalous samples. The best results were obtained when with Support Vectors Machine-Discriminant Analysis (SVM-DA) and Partial Least Squares-Discriminant Analysis (PLS-DA) after mean centering plus multiplicative scatter correction. For SVM-DA the percentage of successful classification was 100% for the training group and 100% and 90% in validation group for non transgenic and transgenic soybean oil samples respectively. For PLS-DA the percentage of successful classification was 95% and 100% in training group for non transgenic and transgenic soybean oil samples respectively and 100% and 80% in validation group for non transgenic and transgenic respectively. The results demonstrate that NIR spectroscopy can provide a rapid, nondestructive and reliable method to distinguish non-transgenic and transgenic soybean oils.

  20. 7 CFR 1437.402 - Carrying capacity.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 7 Agriculture 10 2010-01-01 2010-01-01 false Carrying capacity. 1437.402 Section 1437.402... have a positive impact on the forage's carrying capacity in the crop year NAP assistance is requested... and such practices can be expected to have a positive impact on the forage's carrying capacity in...

  1. 7 CFR 1437.402 - Carrying capacity.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 7 Agriculture 10 2014-01-01 2014-01-01 false Carrying capacity. 1437.402 Section 1437.402... Determining Coverage of Forage Intended for Animal Consumption § 1437.402 Carrying capacity. (a) CCC will establish a carrying capacity for all grazed forage present in the county for purposes of administering...

  2. 7 CFR 1437.402 - Carrying capacity.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 7 Agriculture 10 2013-01-01 2013-01-01 false Carrying capacity. 1437.402 Section 1437.402... Determining Coverage of Forage Intended for Animal Consumption § 1437.402 Carrying capacity. (a) CCC will establish a carrying capacity for all grazed forage present in the county for purposes of administering...

  3. 7 CFR 1437.402 - Carrying capacity.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 7 Agriculture 10 2012-01-01 2012-01-01 false Carrying capacity. 1437.402 Section 1437.402... Determining Coverage of Forage Intended for Animal Consumption § 1437.402 Carrying capacity. (a) CCC will establish a carrying capacity for all grazed forage present in the county for purposes of administering...

  4. Generation of transgenic Hydra by embryo microinjection.

    PubMed

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

    2014-09-11

    As a member of the phylum Cnidaria, the sister group to all bilaterians, Hydra can shed light on fundamental biological processes shared among multicellular animals. Hydra is used as a model for the study of regeneration, pattern formation, and stem cells. However, research efforts have been hampered by lack of a reliable method for gene perturbations to study molecular function. The development of transgenic methods has revitalized the study of Hydra biology(1). Transgenic Hydra allow for the tracking of live cells, sorting to yield pure cell populations for biochemical analysis, manipulation of gene function by knockdown and over-expression, and analysis of promoter function. Plasmid DNA injected into early stage embryos randomly integrates into the genome early in development. This results in hatchlings that express transgenes in patches of tissue in one or more of the three lineages (ectodermal epithelial, endodermal epithelial, or interstitial). The success rate of obtaining a hatchling with transgenic tissue is between 10% and 20%. Asexual propagation of the transgenic hatchling is used to establish a uniformly transgenic line in a particular lineage. Generating transgenic Hydra is surprisingly simple and robust, and here we describe a protocol that can be easily implemented at low cost.

  5. Generation of Transgenic Hydra by Embryo Microinjection

    PubMed Central

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

    2014-01-01

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

  6. 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...

  7. 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. PMID:27081361

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

    PubMed Central

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

    2014-01-01

    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. PMID:27081361

  9. Carry it on the bad side!

    PubMed

    Tan, V; Klotz, M J; Greenwald, A S; Steinberg, M E

    1998-10-01

    Patients with diseased hips often must carry objects while walking, yet they are rarely instructed which hand to use because little has been published on the subject. We sought to evaluate the situation mathematically by determining the hip forces that result when a load is carried in the ipsilateral versus the contralateral hand. Using a free-body diagram of single-leg supported stance, we found that when a load was carried in the contralateral hand, the resultant forces on the hip were increased considerably. Conversely, when the weight was carried in the ipsilateral hand, the forces were actually lower than when no weight was carried at all. Thus, carrying a weight on the opposite side resulted in hip forces that were substantially greater than when the weight was carried on the same side. PMID:9796709

  10. [Transgenic technology and soybean quality improvement].

    PubMed

    Cheng, Hao; Jin, Hang-Xia; Gai, Jun-Yi; Yu, De-Yue

    2011-05-01

    Soybean is an important source of edible oil, protein and protein diet. The breeding process of high quality soybean can be accelerated via employment of transgenic technology, by which the key genes for soybean quality traits could be directly manipulated. Thus, various soybean varieties could be bred to fulfill different needs for specific consumers. Here, we reviewed the contribution of transgenic technology to improvement of soybean qualities in recent years. We also introduce some newly developed safe transgenic technologies and hope this information could relieve some concerns on the GM food.

  11. 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.

  12. 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.

  13. Transgene integration and organization in cotton (Gossypium hirsutum L.) genome.

    PubMed

    Zhang, Jun; Cai, Lin; Cheng, Jiaqin; Mao, Huizhu; Fan, Xiaoping; Meng, Zhaohong; Chan, Ka Man; Zhang, Huijun; Qi, Jianfei; Ji, Lianghui; Hong, Yan

    2008-04-01

    While genetically modified upland cotton (Gossypium hirsutum L.) varieties are ranked among the most successful genetically modified organisms (GMO), there is little knowledge on transgene integration in the cotton genome, partly because of the difficulty in obtaining large numbers of transgenic plants. In this study, we analyzed 139 independently derived T0 transgenic cotton plants transformed by Agrobacterium tumefaciens strain AGL1 carrying a binary plasmid pPZP-GFP. It was found by PCR that as many as 31% of the plants had integration of vector backbone sequences. Of the 110 plants with good genomic Southern blot results, 37% had integration of a single T-DNA, 24% had two T-DNA copies and 39% had three or more copies. Multiple copies of the T-DNA existed either as repeats in complex loci or unlinked loci. Our further analysis of two T1 populations showed that segregants with a single T-DNA and no vector sequence could be obtained from T0 plants having multiple T-DNA copies and vector sequence. Out of the 57 T-DNA/T-DNA junctions cloned from complex loci, 27 had canonical T-DNA tandem repeats, the rest (30) had deletions to T-DNAs or had inclusion of vector sequences. Overlapping micro-homology was present for most of the T-DNA/T-DNA junctions (38/57). Right border (RB) ends of the T-DNA were precise while most left border (LB) ends (64%) had truncations to internal border sequences. Sequencing of collinear vector integration outside LB in 33 plants gave evidence that collinear vector sequence was determined in agrobacterium culture. Among the 130 plants with characterized flanking sequences, 12% had the transgene integrated into coding sequences, 12% into repetitive sequences, 7% into rDNAs. Interestingly, 7% had the transgene integrated into chloroplast derived sequences. Nucleotide sequence comparison of target sites in cotton genome before and after T-DNA integration revealed overlapping microhomology between target sites and the T-DNA (8/8), deletions to

  14. Determination of the expression of fish antifreeze protein (AFP) in 7th generation transgenic mice tissues and serum.

    PubMed

    Bagis, Haydar; Tas, Arzu; Kankavi, Orhan

    2008-06-01

    In this study, the presence of antifreeze protein (AFP) gene expression through successive generations in transgenic mice carrying the chimeric gene construct of the coding sequence for the AFP protein from ocean pout was investigated. AFP transgenic hemizygote mice were used for AFP gene expression. AFP genome expressions in transgenic mice were analyzed by Western blotting, and tissue location of AFP protein was shown by immunohistochemical and immunofluorescence techniques. Seventh transgenic mice from the established founders demonstrated the expression of AFP in organs such as the skin, oviduct, lung, kidney and liver tissues and serum except for the heart. Our results demonstrate successful expression of AFP gene products in several tissues and serum of transgenic mice, the association of in vivo expressed AFP protein, for the first time. These results indicate that the coding sequence for the AFP protein gene (ocean pout type III AFP gene) could be integrated and stably transcribed and expressed in the 7th generation of transgenic mice. In conclusion transgenic mouse lines would be a good model for the cryostudy of AFP and for the determination of AFP roles in several organs and tissues.

  15. Testis-specific expression of a metallothionein I-driven transgene correlates with undermethylation of the locus in testicular DNA.

    PubMed Central

    Salehi-Ashtiani, K; Widrow, R J; Markert, C L; Goldberg, E

    1993-01-01

    Mice carrying a chimeric transgene of the human testis-specific lactate dehydrogenase cDNA driven by mouse metallothionein I promoter have been reported to express the transgene in a testis-specific manner in six founder lines. To study the mechanism by which this testis-specific expression is mediated, we have examined genomic placement, expression pattern, and methylation status of the transgene. Our results indicate that transgene expression is repressed in all somatic tissues examined even when heavy metals are administered. Nuclear run-on assays indicate that failure of expression in the liver (in which the metallothionein I promoter is highly active) occurs at the transcriptional level. In contrast, the transgene mRNA is transcribed in male germ cells and is developmentally regulated during spermatogenesis. Examination of the transgene methylation status reveals that expression is inversely correlated with hypermethylation of the locus; all CpG dinucleotides examined in the promoter region were found to be fully methylated in kidney and liver but were undermethylated in testis. Since methylation of the murine metallothionein I promoter is sufficient to inhibit its activity, it is likely that suppression of the transgene in somatic tissues is mediated by methylation. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 PMID:8415626

  16. [Review of transgenic crop breeding in China].

    PubMed

    Huang, Dafang

    2015-06-01

    The development history and fundamental experience of transgenic crops (Genetically modified crops) breeding in China for near 30 years were reviewed. It was illustrated that a scientific research, development and industrialization system of transgenic crops including gene discovery, transformation, variety breeding, commercialization, application and biosafety assessment has been initially established which was few in number in the world. The research innovative capacity of transgenic cotton, rice and corn has been lifted. The research features as well as relative advantages have been initially formed. The problems and challenges of transgenic crop development were discussed. In addition, three suggestions of promoting commercialization, speeding up implementation of the Major National Project of GM Crops, and enhancing science communication were made. PMID:26672365

  17. 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...

  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. 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.

  20. 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.

  1. Pyramiding transgenes for multiple resistance in rice against bacterial blight, yellow stem borer and sheath blight.

    PubMed

    Datta, K; Baisakh, N; Thet, K Maung; Tu, J; Datta, S K

    2002-12-01

    Here we describe the development of transgene-pyramided stable elite rice lines resistant to disease and insect pests by conventional crossing of two transgenic parental lines transformed independently with different genes. The Xa21 gene (resistance to bacterial blight), the Bt fusion gene (for insect resistance) and the chitinase gene (for tolerance of sheath blight) were combined in a single rice line by reciprocal crossing of two transgenic homozygous IR72 lines. F4 plant lines carrying all the genes of interest stably were identified using molecular methods. The identified lines, when exposed to infection caused by Xanthomonas oryzae pv oryzae, showed resistance to bacterial blight. Neonate larval mortality rates of yellow stem borer ( Scirpophaga incertulas) in an insect bioassay of the same identified lines were 100%. The identified line pyramided with different genes to protect against yield loss showed high tolerance of sheath blight disease caused by Rhizoctonia solani.

  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. Health considerations regarding horizontal transfer of microbial transgenes present in genetically modified crops.

    PubMed

    Kleter, Gijs A; Peijnenburg, Ad A C M; Aarts, Henk J M

    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.

  5. Regulated tissue-specific alternative splicing of enhanced green fluorescent protein transgenes conferred by alpha-tropomyosin regulatory elements in transgenic mice.

    PubMed

    Ellis, Peter D; Smith, Christopher W J; Kemp, Paul

    2004-08-27

    The mutually exclusive exons 2 and 3 of alpha-tropomyosin (alphaTM) have been used as a model system for strictly regulated alternative splicing. Exon 2 inclusion is only observed at high levels in smooth muscle (SM) tissues, whereas striated muscle and non-muscle cells use predominantly exon 3. Experiments in cell culture have shown that exon 2 selection results from repression of exon 3 and that this repression is mediated by regulatory elements flanking exon 3. We have now tested the cell culture-derived model in transgenic mice. We show that by harnessing the intronic splicing regulatory elements, expression of an enhanced green fluorescent protein transgene with a constitutively active promoter can be restricted to SM cells. Splicing of both endogenous alphaTM and a series of transgenes carrying regulatory element mutations was analyzed by reverse transcriptasePCR. These studies indicated that although SM-rich tissues are equipped to regulate splicing of high levels of endogenous or transgene alphaTM RNA, other non-SM tissues such as spleen, which express lower amounts of alphaTM, also splice significant proportions of exon 2, and this splicing pattern can be recapitulated by transgenes expressed at low levels. We confirm the importance in vivo of the negatively acting regulatory elements for regulated skipping of exon 3. Moreover, we provide evidence that some of the regulatory factors responsible for exon 3 skipping appear to be titratable, with loss of regulated splicing sometimes being associated with high transgene expression levels. PMID:15194683

  6. 7 CFR 1437.402 - Carrying capacity.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... factors, such as soil type, elevation, and topography, result in a significant difference of carrying... management and maintenance practices are improvements over those practices generally associated with...

  7. 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.

  8. 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.

  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. PMID:15639876

  10. 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

  11. Transgene flow: facts, speculations and possible countermeasures.

    PubMed

    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

  12. Over-expression of OsHsfA7 enhanced salt and drought tolerance in transgenic rice.

    PubMed

    Liu, Ai-Ling; Zou, Jie; Liu, Cui-Fang; Zhou, Xiao-Yun; Zhang, Xian-Wen; Luo, Guang-Yu; Chen, Xin-Bo

    2013-01-01

    Heat shock proteins play an important role in plant stress tolerance and are mainly regulated by heat shock transcription factors (Hsfs). In this study, we generated transgenic rice over-expressing OsHsfA7 and carried out morphological observation and stress tolerance assays. Transgenic plants exhibited less, shorter lateral roots and root hair. Under salt treatment, over-expressing OsHsfA7 rice showed alleviative appearance of damage symptoms and higher survival rate, leaf electrical conductivity and malondialdehyde content of transgenic plants were lower than those of wild type plants. Meanwhile, transgenic rice seedlings restored normal growth but wild type plants could not be rescued after drought and re-watering treatment. These findings indicate that over-expression of OsHsfA7 gene can increase tolerance to salt and drought stresses in rice seedlings.

  13. Generation and characterization of a transgenic zebrafish expressing the reverse tetracycline transactivator.

    PubMed

    Gu, Qilin; Yang, Xiaojie; He, Xiaozhen; Li, Qing; Cui, Zongbin

    2013-10-20

    Conditional expression of a target gene during zebrafish development is a powerful approach to elucidate gene functions. The tetracycline-controlled systems have been successfully used in the modulation of gene expression in mammalian cells, but few lines of zebrafish carrying these systems are currently available. In this study, we had generated a stable transgenic zebrafish line that ubiquitously expressed the second-generation of reverse Tet transactivator (rtTA-M2). Southern blotting analysis and high-throughput genome sequencing verified that a single copy of rtTA-M2 gene had stably integrated into the zebrafish genome. After induction with doxycycline (Dox), a strong green fluorescent protein (GFP) was seen in rtTA-transgenic eggs injected with pTRE-EGFP plasmids. The fluorescent signal gradually decreased after the withdrawal of Dox and disappeared. However, leaky expression of GFP was undetectable before Dox-induction. Additionally, transgenic embryos expressing rtTA-M2 exhibited no obvious defects in morphological phenotypes, hatching behavior and expression patterns of developmental marker genes, suggesting that rtTA-M2 had little effect on the development of transgenic zebrafish. Moreover, expressed Dickkopf-1 (DKK1) in pTRE-DKK1-injected embryos led to alterations in the expression of marker genes associated with Wnt signaling. Thus, this rtTA-transgenic zebrafish can be utilized to dissect functions of genes in a temporal manner.

  14. Expression of the whey acidic protein in transgenic pigs impairs mammary development.

    PubMed

    Shamay, A; Pursel, V G; Wilkinson, E; Wall, R J; Hennighausen, L

    1992-05-01

    The whey acidic protein has been found in milk of mice, rats, rabbits and camels, and its gene is expressed specifically in mammary tissue at late pregnancy and throughout lactation. A characteristic of whey acidic protein is the 'four-disulfide-core' signature which is also present in proteins involved in organ development. We have generated six lines of transgenic pigs which carry a mouse whey acidic protein transgene and express it at high levels in their mammary glands. Transgenic sows from three lines could not produce sufficient quantities of milk to support normal development of healthy offspring. This phenotype appears to be similar, if not identical, to the milchlos phenotype exhibited by mice expressing whey acidic protein transgenes. Mammary tissue from post-partum milchlos sows had an immature histological appearance, which was distinct from that observed during normal development or involution. Expression of the whey acidic protein transgene was found in mammary tissue from sexually immature pigs from milchlos lines, but not in sows from lines that appeared to lactate normally. We suggest that precocious synthesis of whey acidic protein impairs mammary development and function. Impaired mammary development due to inappropriate timing of whey acidic protein expression is consistent with the notion that proteins with the 'four-disulfide-core' signature participate in tissue formation. PMID:1284481

  15. Autoimmune diabetes can be induced in transgenic major histocompatibility complex class II-deficient mice

    PubMed Central

    1993-01-01

    Insulin-dependent diabetes mellitus (IDDM) is an autoimmune disease marked by hyperglycemia and mononuclear cell infiltration of insulin- producing beta islet cells. Predisposition to IDDM in humans has been linked to the class II major histocompatibility complex (MHC), and islet cells often become aberrantly class II positive during the course of the disease. We have used two recently described transgenic lines to investigate the role of class II molecules and CD4+ T cells in the onset of autoimmune insulitis. Mice that are class II deficient secondary to a targeted disruption of the A beta b gene were bred to mice carrying a transgene for the lymphocytic choriomenigitis virus (LCMV) glycoprotein (GP) targeted to the endocrine pancreas. Our results indicate that class II-deficient animals with and without the GP transgene produce a normal cytotoxic T lymphocyte response to whole LCMV. After infection with LCMV, GP-transgenic class II-deficient animals develop hyperglycemia as rapidly as their class II-positive littermates. Histologic examination of tissue sections from GP- transgenic class II-deficient animals reveals lymphocytic infiltrates of the pancreatic islets that are distinguishable from those of their class II-positive littermates only by the absence of infiltrating CD4+ T cells. These results suggest that in this model of autoimmune diabetes, CD4+ T cells and MHC class II molecules are not required for the development of disease. PMID:8101862

  16. 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.

  17. Weapon-Carrying and Youth Violence.

    ERIC Educational Resources Information Center

    Page, Randy M.; Hammermeister, Jon

    1997-01-01

    Reviews the prevalence of weapon-carrying among adolescents, focusing on the reasons why they carry weapons, ways that firearms are obtained, firearms and violence, and the controlling of weapons in schools. Details weapon-security measures and argues for cooperative action among schools, communities, and government. (RJM)

  18. 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 information…

  19. 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

  20. 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…

  1. Polyamines in response to abiotic stress tolerance through transgenic approaches

    PubMed Central

    Pathak, Malabika Roy; Teixeira da Silva, Jaime A; Wani, Shabir H

    2014-01-01

    The distribution, growth, development and productivity of crop plants are greatly affected by various abiotic stresses. Worldwide, sustainable crop productivity is facing major challenges caused by abiotic stresses by reducing the potential yield in crop plants by as much as 70%. Plants can generally adapt to one or more environmental stresses to some extent. Physiological and molecular studies at transcriptional, translational, and transgenic plant levels have shown the pronounced involvement of naturally occurring plant polyamines (PAs), in controlling, conferring, and modulating abiotic stress tolerance in plants. PAs are small, low molecular weight, non-protein polycations at physiological pH, that are present in all living organisms, and that have strong binding capacity to negatively charged DNA, RNA, and different protein molecules. They play an important role in plant growth and development by controlling the cell cycle, acting as cell signaling molecules in modulating plant tolerance to a variety of abiotic stresses. The commonly known PAs, putrescine, spermidine, and spermine tend to accumulate together accompanied by an increase in the activities of their biosynthetic enzymes under a range of environmental stresses. PAs help plants to combat stresses either directly or by mediating a signal transduction pathway, as shown by molecular cloning and expression studies of PA biosynthesis-related genes, knowledge of the functions of PAs, as demonstrated by developmental studies, and through the analysis of transgenic plants carrying PA genes. This review highlights how PAs in higher plants act during environmental stress and how transgenic strategies have improved our understanding of the molecular mechanisms at play. PMID:24710064

  2. Stability of transgenes in long-term micropropagation of plants of transgenic birch (Betula platyphylla).

    PubMed

    Zeng, Fansuo; Qian, Jingjing; Luo, Wei; Zhan, Yaguang; Xin, Ying; Yang, Chuanping

    2010-01-01

    The stability of integration and expression level of transgenes in long-term micropropagation clones of transgenic birch (Betula platyphylla Suk.) was examined. Multiplexed PCR and reverse primer PCR demonstrated stable integration of transgenes into regenerated plants. Expression levels of the bgt and gus genes among shoot plantlets, subcultured 4, 7, 9 and 15 times, were significantly different. The transcriptional expression level of extraneous genes in regenerated plants decreased with increasing subculture number. Transcriptional gene silencing (TGS) occured in regenerated transgenic lines. The silencing rate of GUS in the 5th subculture plants was 22-65%. TGS in regenerated plants could be reactivated with 5-azacytidine (Azac) at 50-200 microM. GUS and BGT protein expression was reactivated in the micropropagated transgenic birch plants when treated with Azac. A decrease in expression level with increasing number of subcultures is thus associated with DNA methylation.

  3. 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.

  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. 25 CFR 167.6 - Carrying capacities.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ...) Carrying capacities shall be stated in terms of sheep units yearlong, in the ratio of horses, mules, and..., goats, cattle, horses, mules, and burros one year of age or older shall be counted against the...

  6. 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 ...

  7. To carry or not to carry--is this the question? Disentangling the carry effect in multi-digit addition.

    PubMed

    Klein, Elise; Moeller, Korbinian; Dressel, Katharina; Domahs, Frank; Wood, Guilherme; Willmes, Klaus; Nuerk, Hans-Christoph

    2010-09-01

    Recent research has suggested addition performance to be determined by both the need for a carry operation and problem size. Nevertheless, it has remained debatable, how these two factors are interrelated. In the current study, this question was pursued by orthogonally manipulating carry and problem size in two-digit addition verification. As the two factors interacted reliably, our results indicate that the carry effect is moderated by number magnitude processing rather than representing a purely procedural, asemantic sequence of processing steps. Moreover, it was found that the carry effect may not be a purely categorical effect but may be driven by continuous characteristics of the sum of the unit digits as well. Since the correct result of a carry problem can only be derived by integrating and updating the magnitudes of tens and units within the place-value structure of the Arabic number system, the present study provides evidence for the idea that decomposed processing of tens and units also transfers to mental arithmetic.

  8. 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.

  9. Growth factor transgenes interactively regulate articular chondrocytes.

    PubMed

    Shi, Shuiliang; Mercer, Scott; Eckert, George J; Trippel, Stephen B

    2013-04-01

    Adult articular chondrocytes lack an effective repair response to correct damage from injury or osteoarthritis. Polypeptide growth factors that stimulate articular chondrocyte proliferation and cartilage matrix synthesis may augment this response. Gene transfer is a promising approach to delivering such factors. Multiple growth factor genes regulate these cell functions, but multiple growth factor gene transfer remains unexplored. We tested the hypothesis that multiple growth factor gene transfer selectively modulates articular chondrocyte proliferation and matrix synthesis. We tested the hypothesis by delivering combinations of the transgenes encoding insulin-like growth factor I (IGF-I), fibroblast growth factor-2 (FGF-2), transforming growth factor beta1 (TGF-β1), bone morphogenetic protein-2 (BMP-2), and bone morphogenetic protien-7 (BMP-7) to articular chondrocytes and measured changes in the production of DNA, glycosaminoglycan, and collagen. The transgenes differentially regulated all these chondrocyte activities. In concert, the transgenes interacted to generate widely divergent responses from the cells. These interactions ranged from inhibitory to synergistic. The transgene pair encoding IGF-I and FGF-2 maximized cell proliferation. The three-transgene group encoding IGF-I, BMP-2, and BMP-7 maximized matrix production and also optimized the balance between cell proliferation and matrix production. These data demonstrate an approach to articular chondrocyte regulation that may be tailored to stimulate specific cell functions, and suggest that certain growth factor gene combinations have potential value for cell-based articular cartilage repair.

  10. Manipulation of glutathione metabolism in transgenic plants.

    PubMed

    Creissen, G; Broadbent, P; Stevens, R; Wellburn, A R; Mullineaux, P

    1996-05-01

    There is clear potential for the genetic manipulation of key enzymes involved in stress metabolism in transgenic plants. However, the data emerging so far from such experiments are equivocal. The detailed analysis of stress responses in progeny of primary transgenics, coupled with comparisons with control transgenic plants that do not contain the GR transgene, allows us to take into account the possible variation in response to stress associated with regeneration of plants from tissue culture. The picture that is now beginning to emerge with respect to the role of GR in stress protection is that, although there are clearly benefits to be had from overexpression of the enzymes, there is no direct correlation between enzyme levels and stress tolerance. It may be that overexpression of the cytosolic isoform (gor2) will prove to be of greater benefit. Furthermore, the types of stresses to which transgenic plants have been exposed in order to assess the consequences of oxidative stress tolerance cannot reproduce those that will experienced in field conditions. Only when plants with higher GR levels and increased glutathione synthesis capacity are grown in field trials will it be possible to make a full assessment of the benefits of engineering plants with altered glutathione metabolism. PMID:8736785

  11. The transgenic animal platform for biopharmaceutical production.

    PubMed

    Bertolini, L R; Meade, H; Lazzarotto, C R; Martins, L T; Tavares, K C; Bertolini, M; Murray, J D

    2016-06-01

    The recombinant production of therapeutic proteins for human diseases is currently the largest source of innovation in the pharmaceutical industry. The market growth has been the driving force on efforts for the development of new therapeutic proteins, in which transgenesis emerges as key component. The use of the transgenic animal platform offers attractive possibilities, residing on the low production costs allied to high productivity and quality of the recombinant proteins. Although many strategies have evolved over the past decades for the generation of transgenic founders, transgenesis in livestock animals generally faces some challenges, mainly due to random transgene integration and control over transgene copy number. But new developments in gene editing with CRISPR/Cas system promises to revolutionize the field for its simplicity and high efficiency. In addition, for the final approval of any given recombinant protein for animal or human use, the production and characterization of bioreactor founders and expression patterns and functionality of the proteins are technical part of the process, which also requires regulatory and administrative decisions, with a large emphasis on biosafety. The approval of two mammary gland-derived recombinant proteins for commercial and clinical use has boosted the interest for more efficient, safer and economic ways to generate transgenic founders to meet the increasing demand for biomedical proteins worldwide. PMID:26820414

  12. 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.

  13. 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. PMID:26212410

  14. Transgenic dissection of HIV genes involved in lymphoid depletion.

    PubMed Central

    Tinkle, B T; Ueda, H; Ngo, L; Luciw, P A; Shaw, K; Rosen, C A; Jay, G

    1997-01-01

    Transgenic mice carrying an HIV provirus, with selective deletion of all three structural genes, developed extensive lymphoid depletion which was detected not only in the spleen and lymph nodes but also in the thymus. Mice with a high level of HIV gene expression developed acute disease which resulted in premature death, and mice with a low level of viral transcripts developed chronic disease with long-term survival. Neither HIV replication nor the envelope glycoprotein (gp120) was required for T cell depletion. Despite abundant viral gene expression early in life, cell death did not become evident until about the time of full lymphoid maturation, suggesting that thymopoiesis was not affected. The more mature T cells in the peripheral lymphoid organs and in the thymic medulla were less sensitive to the apoptotic process than the immature T cells in the thymic cortex. Gradual depletion of the T cell compartment in the peripheral lymphoid organs was intimately accompanied by the reciprocal expansion of the B cell compartment, resulting in the almost complete replacement of T lymphocytes with B immunoblasts in lymph nodes. Unlike T cells, which showed abundant HIV gene expression, B cells did not. The transgenic approach may help identify the HIV nonstructural gene(s) responsible for immune deficiency and help facilitate dissection of its role in inducing apoptosis. PMID:9202054

  15. The food safety of transgenic animals: implications from traditional breeding.

    PubMed

    Berkowitz, D B

    1993-01-01

    The genetic events associated with traditional selection have implications for the food safety of transgenic animals. Selection has been empirical, relying on the use of the best animals for breeding. Molecular techniques are now being used to identify the genes selected and to describe the differences between alleles that are important in selection to improve quantitative traits. The results of such analyses provide background details of the genetic and physiological effects of the traditional selection of animal lines. Examples of the kinds of genes that may be subject to selection are those coding for peptide hormones, steroid metabolic enzymes, the calcium-channel gating protein, and genes of the major histocompatibility complex. Unselected genes, sometimes with undesirable alleles, may be carried along as "hitchhikers" if they are closely linked to the selected gene. In spite of this potential for physiologically dangerous genetic changes in selected animals, hereditary food toxicity has never been associated with a selected line of the common food animals. This is probably because the allowable physiological range of results of selection is limited by the requirement for healthy, productive animals. Based on these limitations, foods from healthy transgenic animals produced for the purpose of herd improvement are likely to be as safe as the foods from the untransformed parental line. Animals are important indicators of their own food safety.

  16. 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.

  17. [Mosaic expression of the lacZ reporter-gene under control of 5'-regulatory sequences of the alpha-S1-casein gene in transgenic mice].

    PubMed

    Serova, I A; Andreeva, L E; Khaĭdarova, N V; Dias, L P; Dvorianchikov, G A; Burkov, I A; Baginskaia, N V

    2009-01-01

    Phenomenon of mosaic expression at cellular level is widely presented in tissues and organs of transgenic animals. The communication is concerned a study of the mosaics in transgenic mice carrying the lacZ reporter-gene under control of the bovine and goat alpha-S1-casein genes with 5'-flanked sequences of various ex-tent: pCLZ1--721bp, pCLZ2-- 2001 bp and pCLZ3 3409 bp constructs. Five transgenic founders were generated by injection of the recombinant DNA into zygotes: pCLZ 1 - N 16, pCLZ2 - N 37 and pCLZ3 N 7, N 36, and N 48. Positive for J3-galactosidase activity cells were detected in lactating mammary glands of all transgenic females, however, distribution of the positive cells was variable. We observed two types of mosaics: clonal or "lobule" type with positive cells filling the whole of the globule or stochastic type with single positive cells scattered over one or different lobules. Two types of mosaics were characteristic of all the transgenic animals, although, females carrying the pCLZ2 transgene showed "lobule" type more often than transgenic animals with the transgenes pCLZ and pCLZ3. It is suggested that the stochastic type of mosaics occurs in the cells at terminal stage of differentiation, whereas the type arises from positive for P-galactosidase proliferating precursors. Analysis of the inheritance of the transgenes in different lines demonstrated that the pCLZl transgene was inserted in the X-chromosome of the founder whereas the other two localized in autosomes. Localization of the pCLZl transgene in the X-chromosome did not influence the mosaicism; it was similar to that of transgenic animals carrying the transgenes in autosomes. Ectopic expression of the reporter-gene was detected in mandibular glands from the offsprings of the founders N 16 and N 37 only, as well as in atrezed follicles in N 37. The weak ectopic expression saggests that the 5 S-flanked regulatory sequences used in the constructs are able to provide perfect tissue

  18. 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.

  19. Toxins for Transgenic Resistance to Hemipteran Pests

    PubMed Central

    Chougule, Nanasaheb P.; Bonning, Bryony C.

    2012-01-01

    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. PMID:22822455

  20. Transgenic Expression of Cyclin-Dependent Kinase 4 Results in Epidermal Hyperplasia, Hypertrophy, and Severe Dermal Fibrosis

    PubMed Central

    Miliani de Marval, Paula L.; Gimenez-Conti, Irma B.; LaCava, Margaret; Martinez, Luis A.; Conti, Claudio J.; Rodriguez-Puebla, Marcelo L.

    2001-01-01

    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 p27Kip1 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. PMID:11438484

  1. 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. PMID:26911430

  2. 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.

  3. Gene-splitting technology: a novel approach for the containment of transgene flow in Nicotiana tabacum.

    PubMed

    Wang, Xu-Jing; Jin, Xi; Dun, Bao-Qing; 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

  4. Gene-splitting technology: a novel approach for the containment of transgene flow in Nicotiana tabacum.

    PubMed

    Wang, Xu-Jing; Jin, Xi; Dun, Bao-Qing; 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.

  5. 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. PMID:27627325

  6. 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.

  7. Arithmetic coding with constrained carry operations

    NASA Astrophysics Data System (ADS)

    Mahfoodh, Abo-Talib; Said, Amir; Yea, Sehoon

    2015-03-01

    Buffer or counter-based techniques are adequate for dealing with carry propagation in software implementations of arithmetic coding, but create problems in hardware implementations due to the difficulty of handling worst-case scenarios, defined by very long propagations. We propose a new technique for constraining the carry propagation, similar to "bit-stuffing," but designed for encoders that generate data as bytes instead of individual bits, and is based on the fact that the encoder and decoder can maintain the same state, and both can identify the situations when it desired to limit carry propagation. The new technique adjusts the coding interval in a way that corresponds to coding an unused data symbol, but selected to minimize overhead. Our experimental results demonstrate that the loss in compression can be made very small using regular precision for arithmetic operations.

  8. Impact of transgenic technologies on functional genomics.

    PubMed

    Shashikant, Cooduvalli S; Ruddle, Frank H

    2003-07-01

    Gene transfer technologies in mammals are the focus of renewed interest owing to the recent emphasis on analyzing gene function in the postgenomic era. Three important developments in this area include transgenics, gene targeting and nuclear transfer or animal cloning. These technological innovations have enhanced our ability to analyze gene function at the level of the whole organism and have provided the means to modify gene expression. This review discusses the origins and current status of transgenic technologies. Various applications and technologies including chromosome engineering, stem cells, gene traps and modification of livestock are presented. The impact of mouse technologies and genomics on functional analyses is also discussed.

  9. Lectin cDNA and transgenic plants derived therefrom

    SciTech Connect

    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.

  10. 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), and how…

  11. 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…

  12. 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.

  13. 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...

  14. Maize transgenes containing zein promoters are regulated by opaque2

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Transgenes have great potential in crop improvement, but relatively little is known about the epistatic interaction of transgenes with the native genes in the genome. Understanding these interactions is critical for predicting the response of transgenes to different genetic backgrounds and environm...

  15. 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...

  16. [Methods of hygromycin B phosphotransferase activity assay in transgenic plant].

    PubMed

    Zhuo, Qin; Yang, Xiaoguang

    2004-07-01

    Hygromycin B phosphotransferase (HPT) is a widely used selectable marker protein of transgenic plant. Detection of its activity is critical to studies on the development of various transgenic plants, silence of inserted gene, marker-free system development and safety assessment of transgenic food. In this paper, several methods for detecting the activity of this enzyme were reviewed.

  17. 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

  18. 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.

  19. 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.

  20. 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.

  1. 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.

  2. 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.

  3. Inducible gene expression in transgenic Xenopus embryos.

    PubMed

    Wheeler, G N; Hamilton, F S; Hoppler, S

    2000-07-13

    The amphibian Xenopus laevis has been successfully used for many years as a model system for studying vertebrate development. Because of technical limitations, however, molecular investigations have mainly concentrated on early stages. We have developed a straightforward method for stage-specific induction of gene expression in transgenic Xenopus embryos [1] [2]. This method is based on the Xenopus heat shock protein 70 (Xhsp70 [3]) promoter driving the expression of desired gene products. We found that ubiquitous expression of the transgene is induced upon relatively mild heat treatment. Green fluorescent protein (GFP) was used as a marker to monitor successful induction of gene expression in transgenic embryos. We used this method to study the stage specificity of Wnt signalling function. Transient ectopic Wnt-8 expression during early neurulation was sufficient to repress anterior head development and this capacity was restricted to early stages of neurulation. By transient over-expression at different stages of development, we show that frizzled-7 disrupted morphogenesis sequentially from anterior to posterior along the dorsal axis as development proceeds. These results demonstrate that this method for inducible gene expression in transgenic Xenopus embryos will be a very powerful tool for temporal analysis of gene function and for studying molecular mechanisms of vertebrate organogenesis.

  4. 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.

  5. 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.

  6. Effect of transgene number of spontaneous and radiation-induced micronuclei in lacl transgenic mice

    SciTech Connect

    O`Loughlin, K.G.; Hamer, J.D.; Winegar, R.A.; Mirsalis, J.C.; Short, J.M.

    1994-12-31

    Lacl transgenic mice are widely used for the measurement of mutations in specific target issues. The lacl transgene is present in mice as 40 tandem repeats; this sequence is homozygous (contained in both copies of chromosome 5) in C57Bl/6 mice, and is hemizygous in B6C3F1 mice. Previous reports have indicated that tandem repeats can produce chromosome instability, fragile sites, and other effects. To determine whether the presence of the transgene effects micronucleus induction we compared the response of nontransgenic (NTR) to hemizygous (HEMI) transgenic B6C3F1 mice and to hemizygous and homozygous (HOMO) transgenic C57Bl/6 mice. Five mice/group were irradiated with 500 cGy from a {sup 137}Cs source. Bone marrow was harvested 24 hr after treatment and 2000 polychromatic erythrocytes (PCE) were analyzed per animal. The presence or absence of the lacl transgene had no effect in unirradiated mice on the percent of micronucleated PCE (MN) or on the ratio of PCE to total red blood cells for either strain: B6C3F1 mice had MN frequencies of 0.26% and 0.20% for NTR and HEMI mice, respectively; C57Bl/6 mice had MN frequencies of 0.34%, 0.32%, and 0.38% for NTR, HEMI, and HOMO mice, respectively. Radiation-induced micronucleus frequencies were significantly higher in HEMI lacl B6C3F1 mice (2.85%) than in NTR litter mates (1.59%); the converse was true in C57Bl/6 mice: NTR were 2.45%, HEMI were 1.25%, HOMO were 1.65%. These data suggest that the lacl transgene does not cause chromosome instability as measured by spontaneous micronucleus levels. However, the response of these transgenic mice to a variety of clastogenic agents needs to be investigated before they are integrated into standard in vivo assays for chromosome damage.

  7. A decimal carry-free adder

    NASA Astrophysics Data System (ADS)

    Nikmehr, Hooman; Phillips, Braden; Lim, Cheng-Chew

    2005-02-01

    Recently, decimal arithmetic has become attractive in the financial and commercial world including banking, tax calculation, currency conversion, insurance and accounting. Although computers are still carrying out decimal calculation using software libraries and binary floating-point numbers, it is likely that in the near future, all processors will be equipped with units performing decimal operations directly on decimal operands. One critical building block for some complex decimal operations is the decimal carry-free adder. This paper discusses the mathematical framework of the addition, introduces a new signed-digit format for representing decimal numbers and presents an efficient architectural implementation. Delay estimation analysis shows that the adder offers improved performance over earlier designs.

  8. 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.

  9. 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.

  10. 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.

  11. 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.

  12. Severe immunodeficiency associated with a human immunodeficiency virus 1 NEF/3'-long terminal repeat transgene

    PubMed Central

    1994-01-01

    We have generated several transgenic mouse strains carrying a human immunodeficiency virus 1 (HIV-1) NEF/3' long terminal repeat (LTR) transgene under control of a T cell-specific promoter-enhancer element, showing a depletion of CD4+ T cells in the thymus and periphery. The immunological functions of the line with the most dramatic changes in lymphocyte populations, B6/338L, were analyzed in greater detail. The presence of the transgene in the heterozygous animal is associated with a dominant severe immunodeficiency. Older animals develop lymph- adenopathy and splenomegaly. CD4+CD8+ and CD4+CD8- single positive thymocytes already are depleted in these mice at the earliest stages in ontogeny, and peripheral T cells are reduced in frequency and present cell surface marker expression, which is characteristic for memory and activated T cells. The immunological response of B6/338L mice to several viral infections is also greatly impaired. Thus, the HIV-1 NEF/3' LTR as transgene in T cells can cause immunodeficiency and disease with striking similarities to a known retrovirus-induced immunodeficiency called murine AIDS (H. C. Morse III, S. K. Chattopadhyay, M. Makino, T. N. Frederickson, A. W. Hugin, and J. W. Hartley. 1992. AIDS. 6:607). PMID:8113676

  13. Microarray analyses reveal that plant mutagenesis may induce more transcriptomic changes than transgene insertion.

    PubMed

    Batista, Rita; Saibo, Nelson; Lourenço, Tiago; Oliveira, Maria Margarida

    2008-03-01

    Controversy regarding genetically modified (GM) plants and their potential impact on human health contrasts with the tacit acceptance of other plants that were also modified, but not considered as GM products (e.g., varieties raised through conventional breeding such as mutagenesis). What is beyond the phenotype of these improved plants? Should mutagenized plants be treated differently from transgenics? We have evaluated the extent of transcriptome modification occurring during rice improvement through transgenesis versus mutation breeding. We used oligonucleotide microarrays to analyze gene expression in four different pools of four types of rice plants and respective controls: (i) a gamma-irradiated stable mutant, (ii) the M1 generation of a 100-Gy gamma-irradiated plant, (iii) a stable transgenic plant obtained for production of an anticancer antibody, and (iv) the T1 generation of a transgenic plant produced aiming for abiotic stress improvement, and all of the unmodified original genotypes as controls. We found that the improvement of a plant variety through the acquisition of a new desired trait, using either mutagenesis or transgenesis, may cause stress and thus lead to an altered expression of untargeted genes. In all of the cases studied, the observed alteration was more extensive in mutagenized than in transgenic plants. We propose that the safety assessment of improved plant varieties should be carried out on a case-by-case basis and not simply restricted to foods obtained through genetic engineering. PMID:18303117

  14. Dynamics of behavioral disorders in transgenic mice with modeled Alzheimer's disease.

    PubMed

    Semina, I I; Baichurina, A Z; Makarova, E A; Leushina, A V; Kazakevich, Zh V; Gabdrakhmanova, M R; Mukhamed'yarov, M A; Zefirov, A L

    2015-03-01

    Age-related development of behavioral disorders in transgenic mice with modeled Alzheimer's disease carrying V6S3-Tg(APP695)85Dbo Tg(PSENI)85Dbo) genotype was assessed at the age of 7.5, 10 and 20 months in the following tests: open-field, plus maze, T-maze, conditioned passive avoidance response, rotarod, conflict situation with water deprivation, behavioral despair, and arecoline tremor. The main behavioral disorder in transgenic mice at all observation terms was memory impairment in conditioning with positive (but not negative) reinforcement. At the age of 7.5 and 10 months, transgenic mice also showed signs of nonspecific excitement and anxiety, depression-like state, and symptoms of cholinergic deficit. Our results suggest that appropriate age for behavioral tests in studies of effects of potential anti-Alzheimer drugs in transgenic V6S3-Tg(APP695)85Dbo Tg(PSENI)85Dbo) mice is 7.5-10 months.

  15. Altered synaptic plasticity in the mossy fibre pathway of transgenic mice expressing mutant amyloid precursor protein

    PubMed Central

    2010-01-01

    Aβ peptides derived from the cleavage of amyloid precursor protein are widely believed to play an important role in the pathophysiology of Alzheimer's disease. A common way to study the impact of these molecules on CNS function is to compare the physiology of transgenic mice that overproduce Aβ with non-transgenic animals. In the hippocampus, this approach has been frequently applied to the investigation of synaptic transmission and plasticity in the perforant and Schaffer collateral commissural pathways, the first and third components of the classical hippocampal trisynaptic circuit, respectively. Similar studies however have not been carried out on the remaining component of the trisynaptic circuit, the mossy fibre pathway. Using transverse hippocampal slices prepared from ~2 year old animals we have compared mossy fibre synaptic function in wild-type mice and their Tg2576 littermates which age-dependently overproduce Aβ. Input-output curves were not altered in slices from Tg2576 mice, but these animals exhibited a significant loss of the prominent frequency-facilitation expressed by the mossy fibre pathway. In addition to this change in short term synaptic plasticity, high frequency stimulation-induced, NMDA-receptor-independent LTP was absent in slices from the transgenic mice. These data represent the first description of functional deficits in the mossy fibre pathway of Aβ-overproducing transgenic mice. PMID:21040543

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

    PubMed

    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

  17. 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

  18. 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).

  19. Prototypic chromatin insulator cHS4 protects retroviral transgene from silencing in Schistosoma mansoni

    PubMed Central

    Suttiprapa, Sutas; Rinaldi, Gabriel; Brindley, Paul J.

    2011-01-01

    Vesicular stomatitis virus glycoprotein (VSVG) pseudotyped murine leukemia virus (MLV) virions can transduce schistosomes, leading to chromosomal integration of reporter transgenes. To develop VSVG-MLV for functional genomics in schistosomes, the influence of the chicken β-globin cHS4 element, a prototypic chromatin insulator, on transgene expression was examined. Plasmid pLNHX encoding the MLV 5′- and 3′-Long Terminal Repeats (LTRs) flanking the neomycin phosphotransferase gene (neo) was modified to include, within the U3 region of the 3′-LTR, active components of cHS4 insulator, the 250 bp core fused to the 400 bp 3′-region. Cultured larvae of Schistosoma mansoni were transduced with virions from producer cells transfected with control or cHS4-bearing plasmids. Schistosomules transduced with cHS4 virions expressed two to 20 times higher levels of neo than controls, while carrying comparable numbers of integrated proviral transgenes. The findings not only demonstrated that cHS4 was active in schistosomes but also they represent the first report of activity of cHS4 in any Lophotrochozoan species, which has significant implications for evolutionary conservation of heterochromatin regulation. The findings advance prospects for transgenesis in functional genomics of the schistosome genome to discover intervention targets because they provide the means to enhance and extend transgene activity including for vector based RNA interference. PMID:21918820

  20. 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.

  1. Microarray analyses reveal that plant mutagenesis may induce more transcriptomic changes than transgene insertion.

    PubMed

    Batista, Rita; Saibo, Nelson; Lourenço, Tiago; Oliveira, Maria Margarida

    2008-03-01

    Controversy regarding genetically modified (GM) plants and their potential impact on human health contrasts with the tacit acceptance of other plants that were also modified, but not considered as GM products (e.g., varieties raised through conventional breeding such as mutagenesis). What is beyond the phenotype of these improved plants? Should mutagenized plants be treated differently from transgenics? We have evaluated the extent of transcriptome modification occurring during rice improvement through transgenesis versus mutation breeding. We used oligonucleotide microarrays to analyze gene expression in four different pools of four types of rice plants and respective controls: (i) a gamma-irradiated stable mutant, (ii) the M1 generation of a 100-Gy gamma-irradiated plant, (iii) a stable transgenic plant obtained for production of an anticancer antibody, and (iv) the T1 generation of a transgenic plant produced aiming for abiotic stress improvement, and all of the unmodified original genotypes as controls. We found that the improvement of a plant variety through the acquisition of a new desired trait, using either mutagenesis or transgenesis, may cause stress and thus lead to an altered expression of untargeted genes. In all of the cases studied, the observed alteration was more extensive in mutagenized than in transgenic plants. We propose that the safety assessment of improved plant varieties should be carried out on a case-by-case basis and not simply restricted to foods obtained through genetic engineering.

  2. A two-generation reproduction study with transgenic Bt rice TT51 in Wistar rats.

    PubMed

    Wang, Er Hui; Yu, Zhou; Hu, Jing; Jia, Xu Dong; Xu, Hai Bin

    2014-03-01

    TT51 is a transgenic Bt rice created by fusion a synthetic CryAb/CryAc gene into rice MingHui63. A significant number of animal feeding studies with transgenic crops have been carried out with the rapid development of transgenic crops. However, the evidence is far from identifying whether certain novel transgenic crops possess potential danger for human or animal health after long-term consumption. Rice-based diets, containing 60% ordinary grocery rice, MingHui63 rice or TT51 rice by weight, were fed to two generations of male and female rats in order to determine the potential reproductive effects of TT51. In this study, both clinical performance variables and histopathological responses were examined and compared between groups. There were no significant differences between groups on body weights, food consumption, reproductive data and relative organ/body weights. There were some statistically significant differences in hematology and serum chemistry parameters, but no histological abnormalities were seen in the brain, heart, liver, spleen, kidneys, stomach, small intestine, thymus, ovaries, uterus, testes and epididymides. Based on the results, under the circumstance of this study TT51 show no significant differences on reproduction performance of rats compared with MingHui63 and the control.

  3. Can transgenic maize affect soil microbial communities?

    PubMed

    Mulder, Christian; Wouterse, Marja; Raubuch, Markus; Roelofs, Willem; Rutgers, Michiel

    2006-09-29

    The aim of the experiment was to determine if temporal variations of belowground activity reflect the influence of the Cry1Ab protein from transgenic maize on soil bacteria and, hence, on a regulatory change of the microbial community (ability to metabolize sources belonging to different chemical guilds) and/or a change in numerical abundance of their cells. Litter placement is known for its strong influence on the soil decomposer communities. The effects of the addition of crop residues on respiration and catabolic activities of the bacterial community were examined in microcosm experiments. Four cultivars of Zea mays L. of two different isolines (each one including the conventional crop and its Bacillus thuringiensis cultivar) and one control of bulk soil were included in the experimental design. The growth models suggest a dichotomy between soils amended with either conventional or transgenic maize residues. The Cry1Ab protein appeared to influence the composition of the microbial community. The highly enhanced soil respiration observed during the first 72 h after the addition of Bt-maize residues can be interpreted as being related to the presence of the transgenic crop residues. This result was confirmed by agar plate counting, as the averages of the colony-forming units of soils in conventional treatments were about one-third of those treated with transgenic straw. Furthermore, the addition of Bt-maize appeared to induce increased microbial consumption of carbohydrates in BIOLOG EcoPlates. Three weeks after the addition of maize residues to the soils, no differences between the consumption rate of specific chemical guilds by bacteria in soils amended with transgenic maize and bacteria in soils amended with conventional maize were detectable. Reaped crop residues, comparable to post-harvest maize straw (a common practice in current agriculture), rapidly influence the soil bacterial cells at a functional level. Overall, these data support the existence of short

  4. Tissue-specific and developmentally regulated expression of a chimeric actin-globin gene in transgenic mice.

    PubMed Central

    Shani, M

    1986-01-01

    A chimeric plasmid containing about 2/3 of the rat skeletal muscle actin gene plus 730 base pairs of its 5' flanking sequences fused to the 3' end of a human embryonic globin gene (D. Melloul, B. Aloni, J. Calvo, D. Yaffe, and U. Nudel, EMBO J. 3:983-990, 1984) was inserted into mice by microinjection into fertilized eggs. Eleven transgenic mice carrying the chimeric gene with or without plasmid pBR322 DNA sequences were identified. The majority of these mice transmitted the injected DNA to about 50% of their progeny. However, in transgenic mouse CV1, transmission to progeny was associated with amplification or deletion of the injected DNA sequences, while in transgenic mouse CV4 transmission was distorted, probably as a result of insertional mutagenesis. Tissue-specific expression was dependent on the removal of the vector DNA sequences from the chimeric gene sequences prior to microinjection. None of the transgenic mice carrying the chimeric gene together with plasmid pBR322 sequences expressed the introduced gene in striated muscles. In contrast, the six transgenic mice carrying the chimeric gene sequences alone expressed the inserted gene specifically in skeletal and cardiac muscles. Moreover, expression of the chimeric gene was not only tissue specific, but also developmentally regulated. Similar to the endogenous skeletal muscle actin gene, the chimeric gene was expressed at a relatively high level in cardiac muscle of neonatal mice and at a significantly lower level in adult cardiac muscle. These results indicate that the injected DNA included sufficient cis-acting control elements for its tissue-specific and developmentally regulated expression in transgenic mice. Images PMID:3023942

  5. [Animal welfare problems concerning the use of transgenic animals

    PubMed

    Mani, Peter

    1998-01-01

    Using transgenic animals as clinical models pose certain problems since they can suffer. Yet in single cases transgenic animals can reduce the suffering of (other) animals. The permission to generate transgenic animals is not yet clearly regulated in Switzerland. The term "dignity of creature", as formulated in the Swiss Constitution, has to be defined for the Swiss animal protection law. We present the recommendations of the commission for ethical questions concerning transgenic animals appointed by the Federal Council. Partly, these recommendations shall also be applied to the traditional breeding methods. We support the nomination of a national ethics committee for transgenic animals.

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

    PubMed

    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

  7. Collective navigation of cargo-carrying swarms

    PubMed Central

    Shklarsh, Adi; Finkelshtein, Alin; Ariel, Gil; Kalisman, Oren; Ingham, Colin; Ben-Jacob, Eshel

    2012-01-01

    Much effort has been devoted to the study of swarming and collective navigation of micro-organisms, insects, fish, birds and other organisms, as well as multi-agent simulations and to the study of real robots. It is well known that insect swarms can carry cargo. The studies here are motivated by a less well-known phenomenon: cargo transport by bacteria swarms. We begin with a concise review of how bacteria swarms carry natural, micrometre-scale objects larger than the bacteria (e.g. fungal spores) as well as man-made beads and capsules (for drug delivery). A comparison of the trajectories of virtual beads in simulations (using different putative coupling between the virtual beads and the bacteria) with the observed trajectories of transported fungal spores implies the existence of adaptable coupling. Motivated by these observations, we devised new, multi-agent-based studies of cargo transport by agent swarms. As a first step, we extended previous modelling of collective navigation of simple bacteria-inspired agents in complex terrain, using three putative models of agent–cargo coupling. We found that cargo-carrying swarms can navigate efficiently in a complex landscape. We further investigated how the stability, elasticity and other features of agent–cargo bonds influence the collective motion and the transport of the cargo, and found sharp phase shifts and dual successful strategies for cargo delivery. Further understanding of such mechanisms may provide valuable clues to understand cargo-transport by smart swarms of other organisms as well as by man-made swarming robots. PMID:24312731

  8. Collective navigation of cargo-carrying swarms.

    PubMed

    Shklarsh, Adi; Finkelshtein, Alin; Ariel, Gil; Kalisman, Oren; Ingham, Colin; Ben-Jacob, Eshel

    2012-12-01

    Much effort has been devoted to the study of swarming and collective navigation of micro-organisms, insects, fish, birds and other organisms, as well as multi-agent simulations and to the study of real robots. It is well known that insect swarms can carry cargo. The studies here are motivated by a less well-known phenomenon: cargo transport by bacteria swarms. We begin with a concise review of how bacteria swarms carry natural, micrometre-scale objects larger than the bacteria (e.g. fungal spores) as well as man-made beads and capsules (for drug delivery). A comparison of the trajectories of virtual beads in simulations (using different putative coupling between the virtual beads and the bacteria) with the observed trajectories of transported fungal spores implies the existence of adaptable coupling. Motivated by these observations, we devised new, multi-agent-based studies of cargo transport by agent swarms. As a first step, we extended previous modelling of collective navigation of simple bacteria-inspired agents in complex terrain, using three putative models of agent-cargo coupling. We found that cargo-carrying swarms can navigate efficiently in a complex landscape. We further investigated how the stability, elasticity and other features of agent-cargo bonds influence the collective motion and the transport of the cargo, and found sharp phase shifts and dual successful strategies for cargo delivery. Further understanding of such mechanisms may provide valuable clues to understand cargo-transport by smart swarms of other organisms as well as by man-made swarming robots. PMID:24312731

  9. Spread of Plasmids Carrying Multiple GES Variants.

    PubMed

    Cuzon, Gaelle; Bogaerts, Pierre; Bauraing, Caroline; Huang, Te-Din; Bonnin, Rémy A; Glupczynski, Youri; Naas, Thierry

    2016-08-01

    Five GES-producing Enterobacteriaceae isolates that displayed an extended-spectrum β-lactamase (ESBL) phenotype harbored two GES variants: GES-7 ESBL and GES-6 carbapenemase. In all isolates, the two GES alleles were located on the same integron that was inserted into an 80-kb IncM1 self-conjugative plasmid. Whole-genome sequencing suggested in vivo horizontal gene transfer of the plasmid along with clonal diffusion of Enterobacter cloacae To our knowledge, this is the first description in Europe of clustered Enterobacteriaceae isolates carrying two GES β-lactamases, of which one has extended activity toward carbapenems. PMID:27216071

  10. 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.

  11. Cost of carrying out clinical diagnostic tests.

    PubMed Central

    Barnard, D J; Bingle, J P; Garratt, C J

    1978-01-01

    The total cost of performing diagnostic tests in a hospital laboratory during one year was assessed. The largest single item of expenditure was the cost of the salaries of the technical staff, while the cost of reagents (including radiopharmaceuticals) was relatively small. The total costs of carrying out diagnostic tests are much higher than is often recognised by those who request them. The use of relatively expensive, commercially available assay kits saves time and gives good value for money. It may be worth taking this into account when planning hospital budgets. PMID:647338

  12. 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. PMID:24793822

  13. 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.

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

    PubMed

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

    2016-03-25

    Drought and high salinity are two major abiotic factors that restrict alfalfa productivity. A dehydrin protein, CsLEA, from the desert grass Cleistogenes songorica was transformed into alfalfa (Medicago sativa L.) via Agrobacterium-mediated transformation using the bar gene as a selectable marker, and the drought and salt stress tolerances of the transgenic plants were assessed. Thirty-nine of 119 transformants were positive, as screened by Basta, and further molecularly authenticated using PCR and RT-PCR. Phenotype observations revealed that the transgenic plants grew better than the wild-type (WT) plants after 15d of drought stress and 10d of salt stress: the leaves of WT alfalfa turned yellow, whereas the transgenic alfalfa leaves only wilted; after rewatering, the transgenic plants returned to a normal state, though the WT plants could not be restored. Evaluation of physiologic and biochemical indices during drought and salt stresses showed a relatively lower Na(+) content in the leaves of the transgenic plants, which would reduce toxic ion effects. In addition, the transgenic plants were able to maintain a higher relative water content (RWC), higher shoot biomass, fewer photosystem changes, decreased membrane injury, and a lower level of osmotic stress injury. These results demonstrate that overexpression of the CsLEA gene can enhance the drought and salt tolerance of transgenic alfalfa; in addition, carrying the bar gene in the genome may increase herbicide resistance. PMID:26906624

  15. Impact of the ahas transgene and of herbicides associated with the soybean crop on soil microbial communities.

    PubMed

    Souza, Rosinei Aparecida; Babujia, Letícia Carlos; Silva, Adriana Pereira; de Fátima Guimarães, Maria; Arias, Carlos Arrabal; Hungria, Mariangela

    2013-10-01

    Although Brazil has recently reached the position as the second largest producer of genetically modified soybean [Glycine max (L.) Merr.], there are few reports on the effects of transgenic crops and the associated use of specific herbicides on soil microbial communities, both under the edaphoclimatic conditions in Brazil, and in other producer regions in the southern hemisphere. The aim of this study was to evaluate the effects of transgenic soybean containing the ahas gene conferring resistance to herbicides of the imidazolinone group, and of the herbicides associated with transgenic soybeans on the soil microbial community. Twenty field experiments were carried out during three growing seasons (summer of 2006/2007, short-season of 2007 and summer of 2007/2008), in nine municipalities located in six Brazilian states and in the Federal District. The experiments were conducted using a completely randomized block design with four replicates and three treatments: (1) conventional (non-transgenic) soybean cultivar Conquista with conventional herbicides (bentazone + acifluorfen-sodium and other herbicides, depending on the level of infestation in each region); (2) near-isogenic transgenic Cultivance (CV127) containing the ahas gene, with conventional herbicides; (3) transgenic Cultivance with specific herbicide of the imidazolinone group (imazapyr). As the objective of the study was to verify impacts of the transgene and herbicides on the soil microbial community of the whole area and not only a punctual rhizospheric effects, samples were taken at the 0-10 cm layer prior to cropping and at R2 soybean growth stage, between plant rows. Quantitative (microbial biomass C and N, MB-C and MB-N) and qualitative (DGGE of the 16S rDNA region) parameters of soil microbial community were evaluated. No qualitative or quantitative differences were found that could be attributed to the transgene ahas. A comparison of Cultivance soybean with conventional and imidazolinone

  16. Expression of rice thaumatin-like protein gene in transgenic banana plants enhances resistance to fusarium wilt.

    PubMed

    Mahdavi, F; Sariah, M; Maziah, M

    2012-02-01

    The possibility of controlling Fusarium wilt--caused by Fusarium oxysporum sp. cubensec (race 4)--was investigated by genetic engineering of banana plants for constitutive expression of rice thaumatin-like protein (tlp) gene. Transgene was introduced to cauliflower-like bodies' cluster, induced from meristemic parts of male inflorescences, using particle bombardment with plasmid carrying a rice tlp gene driving by the CaMV 35S promoter. Hygromycin B was used as the selection reagent. The presence and integration of rice tlp gene in genomic DNA confirmed by PCR and Southern blot analyses. RT-PCR revealed the expression of transgene in leaf and root tissues in transformants. Bioassay of transgenic banana plants challenged with Fusarium wilt pathogen showed that expression of TLP enhanced resistance to F. oxysporum sp. cubensec (race 4) compared to control plants.

  17. 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.

  18. 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. PMID:26769836

  19. 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.

  20. Anaemia and resistance to malaria in transgenic mice expressing human tumour necrosis factor.

    PubMed Central

    Taverne, J; Sheikh, N; de Souza, J B; Playfair, J H; Probert, L; Kollias, G

    1994-01-01

    Transgenic mice carrying a modified human tumour necrosis factor (huTNF)/beta-globin gene construct linked to the T-cell-specific locus control region of the human CD2 gene express huTNF in their T cells which is released into the circulation and causes the development of a wasting syndrome. We now report that the mice develop anaemia, probably through enhanced erythrophagocytosis rather than inhibition of reticulocyte production. Thus autologous erythrocytes, as well as sheep erythrocytes, were cleared more rapidly from the circulation of transgenic mice than from littermate controls. By contrast, peritoneal macrophages from transgenic mice were less phagocytic in vitro than cells from controls. They also secreted less murine (mu)TNF when stimulated by either bacterial lipopolysaccharide or toxic malarial antigens. The yields of muTNF approached normal levels, however, when these refractory cells from the transgenic mice were stimulated in the presence of a high concentration of indomethacin, suggesting that the production of muTNF was inhibited by enhanced synthesis of prostaglandins. The parasitaemia of transgenic mice infected with Plasmodium yoelii was about 10-fold less at its peak than in controls, although it followed the same time-course, and the multiplication of P. chabaudi was inhibited to an even greater degree. This control of parasitaemia may also be explained by enhancement of macrophage activity, mediated by huTNF acting on the murine p55 receptor, presumably by increasing the removal of parasites by phagocytosis or their killing by toxic products released by the activated macrophages. These observations suggest that a factor in the anaemia of human malaria may be macrophage activation caused by the secretion of TNF that occurs in this disease. PMID:7959874

  1. Overexpression of defense response genes in transgenic wheat enhances resistance to Fusarium head blight

    PubMed Central

    Mackintosh, Caroline A.; Lewis, Janet; Radmer, Lorien E.; Shin, Sanghyun; Heinen, Shane J.; Smith, Lisa A.; Wyckoff, Meagen N.; Dill-Macky, Ruth; Evans, Conrad K.; Kravchenko, Sasha; Baldridge, Gerald D.; Zeyen, Richard J.

    2006-01-01

    Fusarium head blight (FHB) of wheat, caused by Fusarium graminearum and other Fusarium species, is a major disease problem for wheat production worldwide. To combat this problem, large-scale breeding efforts have been established. Although progress has been made through standard breeding approaches, the level of resistance attained is insufficient to withstand epidemic conditions. Genetic engineering provides an alternative approach to enhance the level of resistance. Many defense response genes are induced in wheat during F. graminearum infection and may play a role in reducing FHB. The objectives of this study were (1) to develop transgenic wheat overexpressing the defense response genes α-1-purothionin, thaumatin-like protein 1 (tlp-1), and β-1,3-glucanase; and (2) to test the resultant transgenic wheat lines against F. graminearum infection under greenhouse and field conditions. Using the wheat cultivar Bobwhite, we developed one, two, and four lines carrying the α-1-purothionin, tlp-1, and β-1,3-glucanase transgenes, respectively, that had statistically significant reductions in FHB severity in greenhouse evaluations. We tested these seven transgenic lines under field conditions for percent FHB disease severity, deoxynivalenol (DON) mycotoxin accumulation, and percent visually scabby kernels (VSK). Six of the seven lines differed from the nontransgenic parental Bobwhite line for at least one of the disease traits. A β-1,3-glucanase transgenic line had enhanced resistance, showing lower FHB severity, DON concentration, and percent VSK compared to Bobwhite. Taken together, the results showed that overexpression of defense response genes in wheat could enhance the FHB resistance in both greenhouse and field conditions. PMID:17103001

  2. Separate elements control DJ and VDJ rearrangement in a transgenic recombination substrate.

    PubMed Central

    Ferrier, P; Krippl, B; Blackwell, T K; Furley, A J; Suh, H; Winoto, A; Cook, W D; Hood, L; Costantini, F; Alt, F W

    1990-01-01

    We describe transgenic mice that carry an antigen receptor gene minilocus comprised of germline T cell receptor (TCR) beta variable gene elements (V, D and J) linked to an immunoglobulin (Ig) C mu constant region gene with or without a DNA segment containing the Ig heavy chain transcriptional enhancer (E mu). Transgenic constructs lacking the E mu-containing segment did not undergo detectable rearrangement in any tissue of six independent transgenic lines. In contrast, transgenic constructs containing this DNA segment underwent rearrangement at high frequency in lymphoid tissues, but not other tissues, of four independent lines. Analyses of purified B and T cells, as well as B and T cell lines, from transgenic animals demonstrated that the E mu-containing segment within the construct allowed partial TCR gene assembly (D to J) in both B and T cells. However, complete TCR gene rearrangement within the construct (V to DJ) occurred only in T cells. Therefore, we have demonstrated elements that can control two separate aspects of TCR beta VDJ rearrangement within this construct. One lies within the E mu-containing DNA segment and represents a dominant, cis-acting element that initiates lymphoid cell-specific D beta to J beta rearrangement; various considerations suggest this activity may be related to that of the E mu element. The second element provides T cell-specific control of complete (V beta to DJ beta) variable region gene assembly; it correlates in activity with expression of the unrearranged V beta segment. Images Fig. 1. Fig. 2. Fig. 4. Fig. 5. PMID:2153073

  3. Transgenic inhibitors of RNA interference in Drosophila.

    PubMed

    Chou, Yu-ting; Tam, Bergin; Linay, Fabien; Lai, Eric C

    2007-01-01

    RNA silencing functions as an adaptive antiviral defense in both plants and animals. In turn, viruses commonly encode suppressors of RNA silencing, which enable them to mount productive infection. These inhibitor proteins may be exploited as reagents with which to probe mechanisms and functions of RNA silencing pathways. In this report, we describe transgenic Drosophila strains that allow inducible expression of the viral RNA silencing inhibitors Flock House virus-B2, Nodamura virus-B2, vaccinia virus-E3L, influenza A virus-NS1 and tombusvirus P19. Some of these, especially the B2 proteins, are effective transgenic inhibitors of double strand RNA-induced gene silencing in flies. On the other hand, none of them is effective against the Drosophila microRNA pathway. Their functional selectivity makes these viral silencing proteins useful reagents with which to study biological functions of the Drosophila RNA interference pathway.

  4. Construction of transgenic Trichoderma koningi with chit42 of Metarhizium anisopliae and analysis of its activity against the Asian corn borer.

    PubMed

    Li, Ying Y; Tang, Jun; Fu, Ke H; Gao, Shi G; Wu, Qiong; Chen, Jie

    2012-01-01

    The chit42 gene was cloned from Metarhizium anisopliae CY1 and was inserted into Trichoderma koningii T30 genome by protoplast transformation. Sixteen transgenic isolates were identified by polymerase chain reaction analysis. The chit42 gene was 1275 bp in length and its coded protein was approximately 42 kDa in size. Semi-quantitative RT-PCR analysis and the measurement of the chitinase activity under induced conditions were conducted. Mortality of the Asian corn borer (Ostrinia furnacalis) was used for assessing efficacy of culture filtrates and conidial suspensions of transgenic Trichoderma strains against the insect. The results indicated that the transgenic Trichoderma strain harboring chit42 gene from Metarhizium anisopliae CY1 showed significant lethal effect on the Asian corn borer larvae. Study on growth inhibition of silkworm (Bombyx mori) larvae was carried out. The transgenic Trichoderma could better hinder the growth and development of the silkworm larvae than the wild-type Trichoderma did. The inhibition to the expression of three genes associated with development and anti-stress response in the mid-gut of the Asian corn borer larvae was more significant in the transcriptional level after larvae were fed with transgenic biomass than with the wild type. Evaluation of inhibition on the growth of maize ear rot pathogens was carried out in vitro test and the transgenic strains kept antagonistic activity against Fusarium verticilloides.

  5. Cryptococcus neoformans carried by Odontomachus bauri ants.

    PubMed

    Jesus, Mariana Santos de; Rodrigues, William Costa; Barbosa, Glaucia; Trilles, Luciana; Wanke, Bodo; Lazéra, Márcia dos Santos; Silva, Manuela da

    2012-06-01

    Cryptococcus neoformans is the most common causative agent of cryptococcosis worldwide. Although this fungus has been isolated from a variety of organic substrates, several studies suggest that hollow trees constitute an important natural niche for C. neoformans. A previously surveyed hollow of a living pink shower tree (Cassia grandis) positive for C. neoformans in the city of Rio de Janeiro, Brazil, was chosen for further investigation. Odontomachus bauri ants (trap-jaw ants) found inside the hollow were collected for evaluation as possible carriers of Cryptococcus spp. Two out of 10 ants were found to carry phenoloxidase-positive colonies identified as C. neoformans molecular types VNI and VNII. The ants may have acted as a mechanical vector of C. neoformans and possibly contributed to the dispersal of the fungi from one substrate to another. To the best of our knowledge, this is the first report on the association of C. neoformans with ants of the genus Odontomachus. PMID:22666855

  6. Cryptococcus neoformans carried by Odontomachus bauri ants.

    PubMed

    Jesus, Mariana Santos de; Rodrigues, William Costa; Barbosa, Glaucia; Trilles, Luciana; Wanke, Bodo; Lazéra, Márcia dos Santos; Silva, Manuela da

    2012-06-01

    Cryptococcus neoformans is the most common causative agent of cryptococcosis worldwide. Although this fungus has been isolated from a variety of organic substrates, several studies suggest that hollow trees constitute an important natural niche for C. neoformans. A previously surveyed hollow of a living pink shower tree (Cassia grandis) positive for C. neoformans in the city of Rio de Janeiro, Brazil, was chosen for further investigation. Odontomachus bauri ants (trap-jaw ants) found inside the hollow were collected for evaluation as possible carriers of Cryptococcus spp. Two out of 10 ants were found to carry phenoloxidase-positive colonies identified as C. neoformans molecular types VNI and VNII. The ants may have acted as a mechanical vector of C. neoformans and possibly contributed to the dispersal of the fungi from one substrate to another. To the best of our knowledge, this is the first report on the association of C. neoformans with ants of the genus Odontomachus.

  7. Biochemical characteristics of human renin expressed in transgenic mice.

    PubMed

    Takaori, K; Kim, S; Fukamizu, A; Sagara, M; Hosoi, M; Katsuki, M; Murakami, K; Yamamoto, K

    1993-01-01

    1. Biochemical properties of human renin expressed in transgenic mice (hRN8-12 mice) carrying the human renin gene (Fukamizu et al. Biochem Biophys Res Commun 1989; 165: 826-32) were examined. The optimum pH of the enzymic activity against human angiotensinogen was 5.5 for both plasma and renal human renin in the hRN8-12 mice. Plasma concentrations of human active and inactive renin in the plasma of hRN8-12 mice were 16.7 +/- 2.8 and 79.9 +/- 14.0 pmol of angiotensin Ih-1 ml-1, respectively, thereby indicating that the predominant form of plasma human renin is the inactive form, as is the case in humans. 2. The molecular masses of plasma human active and inactive renin and renal human active renin in the hRN8-12 mice were estimated to be 46 kDa, 48 kDa and 44 kDa, respectively, as determined by h.p.l.c. on G3,000SW. 3. Human renin in the hRN8-12 mouse kidney was bound to a concanavalin A-Sepharose column, and was eluted with alpha-methyl-D-mannoside, showing that this renin is glycosylated, as is native human renin. 4. Low sodium treatment of the hRN8-12 mice for 2 weeks increased plasma human active renin, renal human renin and renal human renin mRNA levels by 2.6-, 3.8- and 2.8-fold, respectively. Thus, the biosynthesis and secretion of renal human renin in these transgenic mice are obviously stimulated by sodium depletion.

  8. 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. PMID:22749907

  9. 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.

  10. Potential transgenic routes to increase tree biomass.

    PubMed

    Dubouzet, Joseph G; Strabala, Timothy J; Wagner, Armin

    2013-11-01

    Biomass is a prime target for genetic engineering in forestry because increased biomass yield will benefit most downstream applications such as timber, fiber, pulp, paper, and bioenergy production. Transgenesis can increase biomass by improving resource acquisition and product utilization and by enhancing competitive ability for solar energy, water, and mineral nutrients. Transgenes that affect juvenility, winter dormancy, and flowering have been shown to influence biomass as well. Transgenic approaches have increased yield potential by mitigating the adverse effects of prevailing stress factors in the environment. Simultaneous introduction of multiple genes for resistance to various stress factors into trees may help forest trees cope with multiple or changing environments. We propose multi-trait engineering for tree crops, simultaneously deploying multiple independent genes to address a set of genetically uncorrelated traits that are important for crop improvement. This strategy increases the probability of unpredictable (synergistic or detrimental) interactions that may substantially affect the overall phenotype and its long-term performance. The very limited ability to predict the physiological processes that may be impacted by such a strategy requires vigilance and care during implementation. Hence, we recommend close monitoring of the resultant transgenic genotypes in multi-year, multi-location field trials. PMID:24094056

  11. Transgenic approaches to western corn rootworm control.

    PubMed

    Narva, Kenneth E; Siegfried, Blair D; Storer, Nicholas P

    2013-01-01

    The western corn rootworm, Diabrotica virgifera virgifera LeConte (Coleoptera: Chrysomelidae) is a significant corn pest throughout the United States corn belt. Rootworm larvae feed on corn roots causing yield losses and control expenditures that are estimated to exceed US$1 billion annually. Traditional management practices to control rootworms such as chemical insecticides or crop rotation have suffered reduced effectiveness due to the development of physiological and behavioral resistance. Transgenic maize expressing insecticidal proteins are very successful in protecting against rootworm damage and preserving corn yield potential. However, the high rate of grower adoption and early reliance on hybrids expressing a single mode of action and low-dose traits threatens the durability of commercialized transgenic rootworm technology for rootworm control. A summary of current transgenic approaches for rootworm control and the corresponding insect resistance management practices is included. An overview of potential new modes of action based on insecticidal proteins, and especially RNAi targeting mRNA coding for essential insect proteins is provided.

  12. 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

  13. Transgenic mouse model of cutaneous adnexal tumors

    PubMed Central

    Kito, Yusuke; Saigo, Chiemi; Atsushi, Kurabayashi; Mutsuo, Furihata; Tamotsu, Takeuchi

    2014-01-01

    TMEM207 was first characterized as being an important molecule for the invasion activity of gastric signet-ring cell carcinoma cells. In order to unravel the pathological properties of TMEM207, we generated several transgenic mouse lines, designated C57BL/6-Tg (ITF-TMEM207), in which murine TMEM207 was ectopically expressed under a truncated (by ~200 bp) proximal promoter of the murine intestinal trefoil factor (ITF) gene (also known as Tff3). Unexpectedly, a C57BL/6-Tg (ITF-TMEM207) mouse line exhibited a high incidence of spontaneous intradermal tumors with histopathological features that resembled those of various human cutaneous adnexal tumors. These tumors were found in ~14% female and 13% of male 6- to 12-month-old mice. TMEM207 immunoreactivity was found in hair follicle bulge cells in non-tumorous skin, as well as in cutaneous adnexal tumors of the transgenic mouse. The ITF-TMEM207 construct in this line appeared to be inserted to a major satellite repeat sequence at chromosome 2, in which no definite coding molecule was found. In addition, we also observed cutaneous adnexal tumors in three other C57BL/6-Tg (ITF-TMEM207) transgenic mouse lines. We believe that the C57BL/6-Tg (ITF-TMEM207) mouse might be a useful model to understand human cutaneous adnexal tumors. PMID:25305140

  14. Arsenic biotransformation and volatilization in transgenic rice.

    PubMed

    Meng, Xiang-Yan; Qin, Jie; Wang, Li-Hong; Duan, Gui-Lan; Sun, Guo-Xin; Wu, Hui-Lan; Chu, Cheng-Cai; Ling, Hong-Qing; Rosen, Barry P; Zhu, Yong-Guan

    2011-07-01

    • Biotransformation of arsenic includes oxidation, reduction, methylation, and conversion to more complex organic arsenicals. Members of the class of arsenite (As(III)) S-adenosylmethyltransferase enzymes catalyze As(III) methylation to a variety of mono-, di-, and trimethylated species, some of which are less toxic than As(III) itself. However, no methyltransferase gene has been identified in plants. • Here, an arsM gene from the soil bacterium Rhodopseudomonas palustris was expressed in Japonica rice (Oryza sativa) cv Nipponbare, and the transgenic rice produced methylated arsenic species, which were measured by inductively coupled plasma mass spectrometry (ICP-MS) and high-performance liquid chromatography-inductively coupled plasma mass spectrometry (HPLC-ICP-MS). • Both monomethylarsenate (MAs(V)) and dimethylarsenate (DMAs(V)) were detected in the roots and shoots of transgenic rice. After 12 d exposure to As(III), the transgenic rice gave off 10-fold greater volatile arsenicals. • The present study demonstrates that expression of an arsM gene in rice induces arsenic methylation and volatilization, theoretically providing a potential stratagem for phytoremediation. PMID:21517874

  15. Increased Incidence of Squamous Cell Carcinomas in Mastomys natalensis Papillomavirus E6 Transgenic Mice during Two-Stage Skin Carcinogenesis

    PubMed Central

    Helfrich, Iris; Chen, Min; Schmidt, Rainer; Fürstenberger, Gerhard; Kopp-Schneider, Annette; Trick, David; Gröne, Hermann-Josef; zur Hausen, Harald; Rösl, Frank

    2004-01-01

    Papillomaviruses cause certain forms of human cancers, most notably carcinomas of the uterine cervix. In contrast to the well-established involvement of papillomavirus infection in the etiology of cervical carcinomas and in carcinomas of a rare hereditary condition, epidermodysplasia verruciformis, a causative role for cutaneous human papillomavirus types in the development of nonmelanoma skin cancer has not been proven. In order to better understand the functions of individual genes of cutaneous papillomavirus types, we generated transgenic mice carrying oncogene E6 of the Mastomys natalensis papillomavirus (MnPV), which causes keratoacanthomas of the skin in its natural host. In the present study, we demonstrate that under conditions of experimental two-stage skin carcinogenesis, fast-paced squamous cell carcinomas develop in nearly 100% of MnPV E6 transgenic mice in comparison to 10% in their nontransgenic littermates (log rank test; P < 0.0001). Therefore, we conclude that the MnPV E6 transgene favors the malignant progression of chemically induced tumors. Whereas an activating H-ras mutation is a consistent feature in benign and malignant tumors in wild-type mice, the majority of papillomas and keratoacanthomas and all squamous cell carcinomas obtained in MnPV E6 transgenic mice contain nonmutated ras alleles. These results indicate that the development of squamous cell carcinomas in MnPV E6 transgenic mice does not depend on an activated H-ras oncogene. PMID:15078961

  16. 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.

  17. Induction of lactogenesis in transgenic virgin pigs: evidence for gene and integration site-specific hormonal regulation.

    PubMed

    Shamay, A; Pursel, V G; Wall, R J; Hennighausen, L

    1992-02-01

    Five month-old transgenic female pigs from three lines carrying the mouse whey acidic protein (WAP) gene and nontransgenic female littermates were implanted with slow-release estrogen and progesterone pellets. Histological analysis of biopsies taken at the time of implantation and 4 weeks later revealed that mammary alveolar development had occurred upon hormonal stimulation in vivo. beta-Casein and beta-lactoglobulin mRNA was found in all induced animals, and WAP mRNA was detected in two of the three transgenic pigs. Differential hormonal regulation between the transgenes in the three lines and also between endogenous milk protein genes was observed in induced mammary tissue cultured in vitro. In the presence of insulin, hydrocortisone, and PRL, beta-casein and WAP mRNA levels increased in all transgenic pigs. In contrast, beta-lactoglobulin mRNA had reached or exceeded lactational levels in response to the in vivo induction, and no further increase was observed in vitro. This suggests that the regulation of the beta-lactoglobulin gene is distinct from that of beta-casein and WAP. Differences were also observed during pregnancy; whereas beta-lactoglobulin gene expression was induced in early pregnancy, a time when PRL levels are low, WAP mRNA levels increased sharply around parturition. Finally, the observation that hormonal regulation of WAP transgenes greatly differed between the three lines suggests that chromatin surrounding the integration site can modify the response of transcription elements. PMID:1569963

  18. Combined expression of antimicrobial genes (Bbchit1 and LJAMP2) in transgenic poplar enhances resistance to fungal pathogens.

    PubMed

    Huang, Yan; Liu, Hong; Jia, Zhichun; Fang, Qing; Luo, Keming

    2012-10-01

    Populus species are susceptible to infection by microbial pathogens that severely affect their growth and substantially decrease their economic value. In this study, two pathogenesis-related protein genes consisting of Beauveria bassiana chitinase (Bbchit1) and motherwort lipid-transfer protein (LJAMP2) were introduced into Chinese white poplar (Populus tomentosa Carr.) via Agrobacterium-mediated transformation using the hygromycin (hyg) and neomycin phosphotransferase (NPTII) genes as selectable markers, respectively. Polymerase chain reaction analysis confirmed the stable integration of transgenes in the genome of transgenic plants. In vitro assays showed that inhibitory activity against the fungal pathogen Alternaria alternata (Fr.) Keissler was evident from the crude leaf extracts from transgenic plants. Importantly, the double-transgenic plants exhibited significantly higher resistance to the pathogen than either of the single-gene transformants and wild-type plants when inoculated with A. alternata. The level of disease reduction in double-transgenic lines was between 82 and 95%, whereas that of single-gene transformants carrying either LJAMP2 or Bbchit1 was between 65 and 89%. These results indicated that the combined expression of the LJAMP2 and Bbchit-1 genes could significantly enhance resistance to necrotrophic fungal pathogens in poplar.

  19. Transgenic Wheat Expressing a Barley UDP-Glucosyltransferase Detoxifies Deoxynivalenol and Provides High Levels of Resistance to Fusarium graminearum.

    PubMed

    Li, Xin; Shin, Sanghyun; Heinen, Shane; Dill-Macky, Ruth; Berthiller, Franz; Nersesian, Natalya; Clemente, Thomas; McCormick, Susan; Muehlbauer, Gary J

    2015-11-01

    Fusarium head blight (FHB), mainly caused by Fusarium graminearum, is a devastating disease of wheat that results in economic losses worldwide. During infection, F. graminearum produces trichothecene mycotoxins, including deoxynivalenol (DON), that increase fungal virulence and reduce grain quality. Transgenic wheat expressing a barley UDP-glucosyltransferase (HvUGT13248) were developed and evaluated for FHB resistance, DON accumulation, and the ability to metabolize DON to the less toxic DON-3-O-glucoside (D3G). Point-inoculation tests in the greenhouse showed that transgenic wheat carrying HvUGT13248 exhibited significantly higher resistance to disease spread in the spike (type II resistance) compared with nontransformed controls. Two transgenic events displayed complete suppression of disease spread in the spikes. Expression of HvUGT13248 in transgenic wheat rapidly and efficiently conjugated DON to D3G, suggesting that the enzymatic rate of DON detoxification translates to type II resistance. Under field conditions, FHB severity was variable; nonetheless, transgenic events showed significantly less-severe disease phenotypes compared with the nontransformed controls. In addition, a seedling assay demonstrated that the transformed plants had a higher tolerance to DON-inhibited root growth than nontransformed plants. These results demonstrate the utility of detoxifying DON as a FHB control strategy in wheat. PMID:26214711

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

    PubMed

    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. 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

  2. Application of lipid extracts from Solidago canadensis to phytomonitoring of PCB126 in transgenic Arabidopsis plants.

    PubMed

    Shimazu, Sayuri; Ohta, Masaya; Ashida, Hitoshi

    2014-09-01

    The aim of this study is to elucidate the effect of lipid extracts from Solidago canadensis for phytomonitoring of polychlorinated biphenyl (PCB) 126 in the transgenic Arabidopsis plant XgD2V11-6 carrying the recombinant guinea pig (g) aryl hydrocarbon receptor (AhR)-mediated β-glucuronidase (GUS) reporter gene expression system. A lipid extract was prepared from S. canadensis and separated into simple lipid, glycolipid, and phospholipid fractions by silica gel column chromatography. Sterylglucoside (SG), monogalactosyldiacylglycerol (MGDG), digalactosyldiacylglycerol (DGDG), and glucosyl ceramide were found in the glycolipid fraction. When the transgenic Arabidopsis plants were treated with the glycolipid fraction together with PCB126, PCB126-induced GUS activity significantly increased in the whole plant. Moreover, S. canadensis-derived SG, MGDG, and DGDG also significantly increased PCB126-induced GUS activity. These results indicated that glycolipids in S. canadensis enhanced the sensitivity of this monitoring assay. PMID:24530184

  3. Changes in fitness-associated traits due to the stacking of transgenic glyphosate resistance and insect resistance in Brassica napus L.

    PubMed

    Londo, J P; Bollman, M A; Sagers, C L; Lee, E H; Watrud, L S

    2011-10-01

    Increasingly, genetically modified crops are being developed to express multiple 'stacked' traits for different types of transgenes, for example, herbicide resistance, insect resistance, crop quality and tolerance to environmental stresses. The release of crops that express multiple traits could result in ecological changes in weedy environments if feral crop plants or hybrids formed with compatible weeds results in more competitive plants outside of agriculture. To examine the effects of combining transgenes, we developed a stacked line of canola (Brassica napus L.) from a segregating F(2) population that expresses both transgenic glyphosate resistance (CP4 EPSPS) and lepidopteran insect resistance (Cry1Ac). Fitness-associated traits were evaluated between this stacked genotype and five other Brassica genotypes in constructed mesocosm plant communities exposed to insect herbivores (Plutella xylostella L.) or glyphosate-drift. Vegetative biomass, seed production and relative fecundity were all reduced in stacked trait plants when compared with non-transgenic plants in control treatments, indicating potential costs of expressing multiple transgenes without selection pressure. Although costs of the transgenes were offset by selective treatment, the stacked genotype continued to produce fewer seeds than either single transgenic line. However, the increase in fitness of the stacked genotype under selective pressure contributed to an increased number of seeds within the mesocosm community carrying unselected, hitchhiking transgenes. These results demonstrate that the stacking of these transgenes in canola results in fitness costs and benefits that are dependent on the type and strength of selection pressure, and could also contribute to changes in plant communities through hitchhiking of unselected traits.

  4. In situ methods to localize transgenes and transcripts in interphase nuclei: a tool for transgenic plant research

    PubMed Central

    Santos, Ana Paula; Wegel, Eva; Allen, George C; Thompson, William F; Stoger, Eva; Shaw, Peter; Abranches, Rita

    2006-01-01

    Genetic engineering of commercially important crops has become routine in many laboratories. However, the inability to predict where a transgene will integrate and to efficiently select plants with stable levels of transgenic expression remains a limitation of this technology. Fluorescence in situ hybridization (FISH) is a powerful technique that can be used to visualize transgene integration sites and provide a better understanding of transgene behavior. Studies using FISH to characterize transgene integration have focused primarily on metaphase chromosomes, because the number and position of integration sites on the chromosomes are more easily determined at this stage. However gene (and transgene) expression occurs mainly during interphase. In order to accurately predict the activity of a transgene, it is critical to understand its location and dynamics in the three-dimensional interphase nucleus. We and others have developed in situ methods to visualize transgenes (including single copy genes) and their transcripts during interphase from different tissues and plant species. These techniques reduce the time necessary for characterization of transgene integration by eliminating the need for time-consuming segregation analysis, and extend characterization to the interphase nucleus, thus increasing the likelihood of accurate prediction of transgene activity. Furthermore, this approach is useful for studying nuclear organization and the dynamics of genes and chromatin. PMID:17081287

  5. Definition of the human N-myc promoter region during development in a transgenic mouse model.

    PubMed

    Tai, K F; Rogers, S W; Pont-Kingdon, G; Carroll, W L

    1999-09-01

    The N-myc oncogene directs organogenesis, and gene amplification is associated with aggressive forms of neuroblastoma, a common malignant tumor in children. N-myc is expressed in fetal epithelium, and expression decreases markedly postnatally. To localize sequences responsible for directing expression, we have analyzed the human N-myc promoter. We noted previously that N-myc promoter regions 5' to exon 1 directed reporter gene expression in all cell lines, including those without detectable N-myc transcripts. However, when promoter constructs included 3' exon 1 and the 5' portion of intron 1, reporter activity was detected only when there was expression of the endogenous gene. To determine the role of this "tissue-specific region" in directing expression during development, we generated transgenic mice carrying N-myc promoter lacZ minigenes that contained 5' N-myc promoter elements alone or the promoter linked to the 3' exon 1/5' intron 1 tissue-specific region. Animals lacking the tissue-specific exon 1/intron 1 region showed beta-galactosidase expression in the CNS, but expression was not observed in other organs in which endogenously derived N-myc transcripts were seen. Within the CNS, transgene expression was seen mainly in the olfactory system and was not observed in other areas in which expression of the murine gene has been noted. In contrast, no transgene expression was observed in any of the animals carrying the tissue-specific exon 1/intron 1 region. Thus, sequences that direct expression within the olfactory system were contained within our 5' promoter transgene, whereas sequences that guide the ubiquitous expression of N-myc during organogenesis lie outside the regions studied here. Finally, the exon 1/intron 1 region seems to act in a dominant fashion to repress expression in the CNS from the immediate 5' N-myc promoter. PMID:10473038

  6. [Effect of transgenic plants on biodiversity of agroecosystem].

    PubMed

    Nie, Chengrong; Wang, Jianwu; Luo, Shiming

    2003-08-01

    The effect of transgenic plants on the biodiversity of agroecosystem is an important environmental issue. There are many researches in this field at home and abroad recently. This paper reviewed the advances of the researches based on three levels of biodiversity as genetic diversity, species diversity and ecosystem diversity. They included following aspects: the effect of insect-resistant transgenic crops on target pest; the effect of herbicide-resistant transgenic crops on crops and wild weedy relatives; the effect of virus-resistant transgenic crops on virus; and the effect of transgenic crops on non-target organisms. This paper also discussed the effect of transgenic crops on soil ecosystem and crop genetic diversity. Their potential risks included uncontrolled flows of genes to wild relatives; development of herbicide, insect, and virus resistance in wild relatives; reduced crop genetic diversity; and adverse effects on organisms that were not pests, such as beneficial insects.

  7. Transgenic plants as vital components of integrated pest management.

    PubMed

    Kos, Martine; van Loon, Joop J A; Dicke, Marcel; Vet, Louise E M

    2009-11-01

    Although integrated pest management (IPM) strategies have been developed worldwide, further improvement of IPM effectiveness is required. The use of transgenic technology to create insect-resistant plants can offer a solution to the limited availability of highly insect-resistant cultivars. Commercially available insect-resistant transgenic crops show clear benefits for agriculture and there are many exciting new developments such as transgenic plants that enhance biological control. Effective evaluation tools are needed to ascertain that transgenic plants do not result in undesired non-target effects. If these conditions are met, there will be ample opportunities for transgenic plants to become key components of environmentally benign and durable pest management systems. Here we discuss the potential and challenges for incorporating transgenic plants in IPM.

  8. Expression of complete metabolic pathways in transgenic plants.

    PubMed

    Krichevsky, Alexander; Zaltsman, Adi; King, Lisa; Citovsky, Vitaly

    2012-01-01

    Plant genetic engineering emerged as a methodology to introduce only few transgenes into the plant genome. Following fast-paced developments of the past few decades, engineering of much larger numbers of transgenes became a reality, allowing to introduce full metabolic pathways from other organisms into plants and generate transgenics with startling new traits. From the advent of the classical plant genetic engineering, the transgenes were introduced into the nuclear genome of the plant cell, and this strategy still is quite successful when applied to few transgenes. However, for introducing large number of transgenes, we advocate that the chloroplast genome is a superior choice, especially for engineering of new complete metabolic pathways into plants. The ability to genetically engineer plants with complex and fully functional metabolic pathways from other organisms bears a substantial promise in generation of pharmaceuticals, i.e., biopharming, and new agricultural crops with that traits never existed before, leading to enhancement in quality of human life. PMID:22616478

  9. Impact of cry1AC-carrying Brassica rapa subsp. pekinensis on leaf bacterial community.

    PubMed

    Kim, Young Tae; Lee, Kang Seon; Kim, Moon Jung; Kim, Seung Bum

    2009-02-01

    The effects of Chinese cabbage (Brassica rapa subsp. pekinensis) carrying cry1AC derived from Bacillus thuringiensis (Bt) on leaf bacterial community were examined by analyzing the horizontal transfer of trans-gene fragments from plants to bacteria. The effect of plant pathogenic bacteria on the gene transfer was also examined using Pseudomonas syringae pathovar. maculicola. The frequency of hygromycin-resistant bacteria did not alter in Bt leaves, though slight increase was observed in Pseudomonas-infected Bt leaves with no statistical significance. The analysis of bacterial community profiles using the denaturing gradient gel electrophoresis (DGGE) fingerprinting indicated that there were slight differences between Bt and control Chinese cabbage, and also that infected tissues were dominated by P. syringae pv. maculicola. However, the cultured bacterial pools were not found to contain any transgene fragments. Thus, no direct evidence of immediate gene transfer from plant to bacteria or acquisition of hygromycin resistance could be observed. Still, long-term monitoring on the possibility of gene transfer is necessary to correctly assess the environmental effects of the Bt crop on bacteria.

  10. 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

  11. 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

  12. 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

  13. Osteogenic capacity of transgenic flax scaffolds.

    PubMed

    Gredes, Tomasz; Wróbel-Kwiatkowska, Magdalena; Dominiak, Marzena; Gedrange, Tomasz; Kunert-Keil, Christiane

    2012-02-01

    The modification of flax fibers to create biologically active dressings is of undoubted scientific and practical interest. Flax fibers, derived from transgenic flax expressing three bacterial genes for the synthesis of poly-3-hydroxybutyric acid (PHB), have better mechanical properties than unmodified flax fibers; do not show any inflammation response after subcutaneous insertion; and have a good in vitro and in vivo biocompatibility. The aim of this study was to examine the applicability of composites containing flax fibers of genetically modified (M50) or non-modified (wt-Nike) flax within a polylactide (PLA) matrix for bone regeneration. For this, the mRNA expression of genes coding for growth factors (insulin-like growth factor IGF1, IGF2, vascular endothelial growth factor), for osteogenic differentiation (alkaline phosphatase, osteocalcin, Runx2, Phex, type 1 and type 2 collagen), and for bone resorption markers [matrix metalloproteinase 8 (MMP8), acid phosphatase type 5] were analyzed using quantitative real-time polymerase chain reaction. We found a significant elevated mRNA expression of IGF1 with PLA and PLA-wt-Nike composites. The mRNA amount of MMP8 and osteocalcin was significantly decreased in all biocomposite-treated cranial tissue samples compared to controls, whereas the expression of all other tested transcripts did not show any differences. It is assumed that both flax composites are able to stimulate bone regeneration, but composites from transgenic flax plants producing PHB showed faster bone regeneration than composites of non-transgenic flax plants. The application of these linen membranes for bone tissue engineering should be proved in further studies. PMID:22718592

  14. 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

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

    PubMed

    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

  16. 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...

  17. Cloned transgenic heart-healthy pork?

    PubMed

    Prather, Randall S

    2006-08-01

    Here I comment on the production and uses of swine that express a humanized fat-1 gene. The gene product is a fatty acid desaturase that converts omega-6 fatty acids to omega-3 fatty acids. Omega-3 fatty acids have been implicated as being important for reproductive success, maintaining a healthy cardiovascular system, sustaining a functional immune system, and even preventing depression and cancer. The descendants of these hfat-1 transgenic swine will be very useful as models of the human condition, and if they are permitted to enter the food chain, they may improve human health.

  18. 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.

  19. Transgenic production of arachidonic acid in oilseeds.

    PubMed

    Petrie, James R; Shrestha, Pushkar; Belide, Srinivas; Mansour, Maged P; Liu, Qing; Horne, James; Nichols, Peter D; Singh, Surinder P

    2012-02-01

    We describe a transgenic microalgal Δ9-elongase pathway transformed in both Brassica napus and Arabidopsis thaliana seed resulting in the production of arachidonic acid (ARA). This pathway is noteworthy for both the production of ARA in seed tissue and the low levels of intermediate C20 fatty acids that accumulate. We also demonstrate that the arachidonic acid is naturally enriched at the sn2 position in triacylglycerol. This is the first report of ARA production by the Δ9-elongase pathway in an oilseed.

  20. A Transgenic Tri-Modality Reporter Mouse

    PubMed Central

    Yan, Xinrui; Ray, Pritha; Paulmurugan, Ramasamy; Tong, Ricky; Gong, Yongquan; Sathirachinda, Ataya; Wu, Joseph C.; Gambhir, Sanjiv S.

    2013-01-01

    Transgenic mouse with a stably integrated reporter gene(s) can be a valuable resource for obtaining uniformly labeled stem cells, tissues, and organs for various applications. We have generated a transgenic mouse model that ubiquitously expresses a tri-fusion reporter gene (fluc2-tdTomato-ttk) driven by a constitutive chicken β-actin promoter. This “Tri-Modality Reporter Mouse” system allows one to isolate most cells from this donor mouse and image them for bioluminescent (fluc2), fluorescent (tdTomato), and positron emission tomography (PET) (ttk) modalities. Transgenic colonies with different levels of tri-fusion reporter gene expression showed a linear correlation between all three-reporter proteins (R2=0.89 for TdTomato vs Fluc, R2=0.94 for Fluc vs TTK, R2=0.89 for TdTomato vs TTK) in vitro from tissue lysates and in vivo by optical and PET imaging. Mesenchymal stem cells (MSCs) isolated from this transgenics showed high level of reporter gene expression, which linearly correlated with the cell numbers (R2=0.99 for bioluminescence imaging (BLI)). Both BLI (R2=0.93) and micro-PET (R2=0.94) imaging of the subcutaneous implants of Tri-Modality Reporter Mouse derived MSCs in nude mice showed linear correlation with the cell numbers and across different imaging modalities (R2=0.97). Serial imaging of MSCs transplanted to mice with acute myocardial infarction (MI) by intramyocardial injection exhibited significantly higher signals in MI heart at days 2, 3, 4, and 7 (p<0.01). MSCs transplanted to the ischemic hindlimb of nude mice showed significantly higher BLI and PET signals in the first 2 weeks that dropped by 4th week due to poor cell survival. However, laser Doppler perfusion imaging revealed that blood circulation in the ischemic limb was significantly improved in the MSCs transplantation group compared with the control group. In summary, this mouse can be used as a source of donor cells and organs in various research areas such as stem cell research

  1. 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

  2. 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. :

  3. 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.

  4. Cyclin D2 Overexpression in Transgenic Mice Induces Thymic and Epidermal Hyperplasia whereas Cyclin D3 Expression Results Only in Epidermal Hyperplasia

    PubMed Central

    Rodriguez-Puebla, Marcelo L.; LaCava, Margaret; Miliani de Marval, Paula L.; Jorcano, Jose L.; Richie, Ellen R.; Conti, Claudio J.

    2000-01-01

    In a previous report, we described the effects of cyclin D1 expression in epithelial tissues of transgenic mice. To study the involvement of D-type cyclins (D1, D2, and D3) in epithelial growth and differentiation and their putative role as oncogenes in skin, transgenic mice were developed which carry cyclin D2 or D3 genes driven by a keratin 5 promoter. As expected, both transgenic lines showed expression of these proteins in most of the squamous tissues analyzed. Epidermal proliferation increased in transgenic animals and basal cell hyperplasia was observed. All of the animals also had a minor thickening of the epidermis. The pattern of expression of keratin 1 and keratin 5 indicated that epidermal differentiation was not affected. Transgenic K5D2 mice developed mild thymic hyperplasia that reversed at 4 months of age. On the other hand, high expression of cyclin D3 in the thymus did not produce hyperplasia. This model provides in vivo evidence of the action of cyclin D2 and cyclin D3 as mediators of proliferation in squamous epithelial cells. A direct comparison among the three D-type cyclin transgenic mice suggests that cyclin D1 and cyclin D2 have similar roles in epithelial thymus cells. However, overexpression of each D-type cyclin produces a distinct phenotype in thymic epithelial cells. PMID:10980142

  5. 26-Week oral safety study in macaques for transgenic rice containing major human T-cell epitope peptides from Japanese cedar pollen allergens.

    PubMed

    Domon, Eiji; Takagi, Hidenori; Hirose, Sakiko; Sugita, Koichi; Kasahara, Saori; Ebinuma, Hiroyasu; Takaiwa, Fumio

    2009-06-24

    A study of repeated oral administration of transgenic rice containing a hybrid peptide of major human T-cell epitopes (7Crp) from Japanese cedar pollen allergens was carried out in cynomolgus macaques over 26 weeks. The monkeys were divided into three groups, each comprising three males and three females, administered a high dose of transgenic rice, a low dose of transgenic rice, or a high dose of the parental rice strain. The transgenic rice 7crp#10 and the parental nontransgenic control were polished, steamed, mashed, and prepared in water at 40% (w/v). Monkeys were orally administered a high or low dose of transgenic rice or the nontransgenic control by gavage every day. No adverse effects on general behavior or body weight of animals were observed during the study. Analysis of blood from monkeys administered for 26 weeks showed that, with few exceptions, there were no significant differences in hematological or biochemical values between them. Additionally, neither pathological symptoms nor histopathological abnormalities were observed. Thus, it was concluded that oral administration of transgenic rice containing T-cell epitopes from Japanese cedar pollen allergens has no adverse effects.

  6. 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

  7. Efficient and specific ribozyme-mediated reduction of bovine alpha-lactalbumin expression in double transgenic mice.

    PubMed Central

    L'Huillier, P J; Soulier, S; Stinnakre, M G; Lepourry, L; Davis, S R; Mercier, J C; Vilotte, J L

    1996-01-01

    Transgenic mice carrying a bovine alpha-lactalbumin (alpha-lac) specific ribozyme gene under the transcriptional control of the mouse mammary tumor virus long terminal repeat were generated and cross-bred with animals that highly express a bovine alpha-lac transgene (0.4 mg of alpha-lac/ml(-1) of milk). The ribozyme contains the hammerhead catalytic domain, flanked by 12-nt sequences complementary to the 3' untranslated region of bovine alpha-lac transcript. High-level expression of the ribozyme gene was detected by Northern blot analysis in the mammary gland of 7-8 day lactating transgenic mice, from 3 of 12 lines analyzed. Heterozygous expression of the ribozyme resulted in a reduction in the levels of the target mRNA to 78, 58, and 50% of that observed in the nonribozyme transgenic littermate controls for three independent lines. The ribozyme-mediated reduction in the levels of the bovine protein paralleled that observed for the mRNA, and was positively correlated with the level of expression of the ribozyme. In nonribozyme expressing transgenic mice, the level of bovine alpha-lac mRNA and protein was not affected. The specificity of this activity is demonstrated by the absence of a reduction in the levels of the endogenous murine alpha-lac mRNA or protein. These results demonstrate the feasibility of ribozyme-mediated down-regulation of highly-expressed transcripts in transgenic animals. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 Fig. 5 Fig. 6 PMID:8692881

  8. Transgenic plums (Prunus domestica L.) express the plum pox virus coat protein gene.

    PubMed

    Scorza, R; Ravelonandro, M; Callahan, A M; Cordts, J M; Fuchs, M; Dunez, J; Gonsalves, D

    1994-11-01

    Plum hypocotyl slices were transformed with the coat protein (CP) gene of plum pox virus (PPV-CP) following cocultivation with Agrobacterium tumefaciens containing the plasmid pGA482GG/PPVCP-33. This binary vector carries the PPV-CP gene construct, as well as the chimeric neomycin phosphotransferase and β-glucuronidase genes. Integration and expression of the transferred genes into regenerated plum plants was verified through kan resistance, GUS assays, and PCR amplification of the PPV-CP gene. Twenty-two transgenic clones were identified from approximately 1800 hypocotyl slices. DNA, mRNA, and protein analyses of five transgenic plants confirmed the integration of the engineered CP gene, the accumulation of CP mRNA and of PPV-CP-immunoreactive protein. CP mRNA levels ranged from high to undetectable levels, apparently correlated with gene structure, as indicated by DNA blot analysis. Western analysis showed that transgenic plants produced amounts of CP which generally correlated with amounts of detected mRNA.

  9. Generation of transgenic Drosophila expressing shRNAs in the miR-1 backbone.

    PubMed

    Chang, Kenneth; Marran, Krista; Valentine, Amy; Hannon, Gregory J

    2014-05-01

    In Drosophila, long-term effects of RNA interference (RNAi) must be achieved by integrating into the genome a template from which an RNAi trigger is transcribed by cellular RNA polymerases, generally RNA polymerase II or III. With encoded triggers, not only can essentially permanent silencing be achieved, but control can also be exerted over the level of trigger expression, with a resulting variation in the degree to which the target is silenced. Knockdown can also be controlled in a temporal and cell-type-dependent fashion through the use of well-established transgenic methodologies and well-tested promoters. The forms of encoded triggers vary. Long double-stranded RNAs can be expressed as extended inverted repeats. The nearest equivalent of a small interfering RNA is an artificial microRNA (miRNA) or short hairpin RNA (shRNA), where a natural miRNA backbone (also called a scaffold) is remodeled to produce a different small RNA or a small inverted repeat (<30 nucleotides) is simply expressed. This protocol describes creation of transgenic Drosophila carrying shRNA inserts in a remodeled endogenous miRNA backbone. The protocol applies to the use of miRNA-based shRNAs, but most of the vectors, principles of experimental design, and methods are also applicable to long inverted repeat transgenes. PMID:24786506

  10. 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

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

    PubMed Central

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

    2013-01-01

    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 (FISH), 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. PMID:23749207

  12. The development of transgenic mice for the expression of large amounts of human lysozyme in milk.

    PubMed

    Wu, Xiaojie; Lin, Yanli; Xi, Yongyi; Shao, Zhenlu; Zhou, Yanrong; Liu, Fang; Chen, Hongxing

    2014-06-01

    Human lysozyme (hLYZ) has important potential applications as antimicrobial medicine and food additive. To develop a robust expression vector that ensures expression of large amounts of hLYZ in milk, here a 26,267 bp chimeric mouse whey acidic protein (mWAP)::hLYZ cassette was constructed and used as a mammary gland-specific expression vector, in which a 3,010 bp genomic sequence in the 24,466 bp mWAP gene locus was substituted by a 4,811 bp genomic sequence of hLYZ, exactly from the start codon to the stop codon. Corresponding transgenic mice were generated, and enzymatically-active hLYZ was expressed at 18.4-35 g l(-1) in the milk of most transgenic mouse lines. Our transgenic mice carrying chimeric mWAP::hLYZ represent a model system for cost-effective production of hLYZ. PMID:24563307

  13. 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.

  14. 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.

  15. Starch self-processing in transgenic sweet potato roots expressing a hyperthermophilic α-amylase.

    PubMed

    Santa-Maria, Monica C; Yencho, Craig G; Haigler, Candace H; Thompson, William F; Kelly, Robert M; Sosinski, Bryon

    2011-01-01

    Sweet potato is a major crop in the southeastern United States, which requires few inputs and grows well on marginal land. It accumulates large quantities of starch in the storage roots and has been shown to give comparable or superior ethanol yields to corn per cultivated acre in the southeast. Starch conversion to fermentable sugars (i.e., for ethanol production) is carried out at high temperatures and requires the action of thermostable and thermoactive amylolytic enzymes. These enzymes are added to the starch mixture impacting overall process economics. To address this shortcoming, the gene encoding a hyperthermophilic α-amylase from Thermotoga maritima was cloned and expressed in transgenic sweet potato, generated by Agrobacterium tumefaciens-mediated transformation, to create a plant with the ability to self-process starch. No significant enzyme activity could be detected below 40°C, but starch in the transgenic sweet potato storage roots was readily hydrolyzed at 80°C. The transgene did not affect normal storage root formation. The results presented here demonstrate that engineering plants with hyperthermophilic glycoside hydrolases can facilitate cost effective starch conversion to fermentable sugars. Furthermore, the use of sweet potato as an alternative near-term energy crop should be considered.

  16. A plant mitochondrial sequence transcribed in transgenic tobacco chloroplasts is not edited

    SciTech Connect

    Sutton, C.A.; Hanson, M.R.; Zoubenko, O.V.; Maliga, P.

    1995-03-01

    RNA editing occurs in two higher-plant organelles, chloroplasts, and mitochondria. Because chloroplasts and mitochondria exhibit some similarity in editing site selection, we investigated whether mitochondrial RNA sequences could be edited in chloroplasts. We produced transgenic tobacco plants that contained chimeric genes in which the second exon of a Petunia hybrida mitochondrial coxII gene was under the control of chloroplast gene regulatory sequences. coxII transcripts accumulated to low or high levels in transgenic chloroplasts containing chimeric genes with the plastid ribosomal protein gene rps16 or the rRNA operon promoter, respectively. Exon 2 of coxII was chosen because it carries seven editing sites and is edited in petunia mitochondria even when located in an abnormal context in an aberrant recombined gene. When editing of the coxII transcripts in transgenic chloroplasts was examined, no RNA editing at any of the usual sites was detected, nor was there any novel editing at any other sites. These results indicate that the RNA editing mechanisms of chloroplasts and mitochondria are not identical but must have at least some organelle-specific components. 33 refs., 5 figs.

  17. Transgenic plants as factories for biopharmaceuticals.

    PubMed

    Giddings, G; Allison, G; Brooks, D; Carter, A

    2000-11-01

    Plants have considerable potential for the production of biopharmaceutical proteins and peptides because they are easily transformed and provide a cheap source of protein. Several biotechnology companies are now actively developing, field testing, and patenting plant expression systems, while clinical trials are proceeding on the first biopharmaceuticals derived from them. One transgenic plant-derived biopharmaceutical, hirudin, is now being commercially produced in Canada for the first time. Product purification is potentially an expensive process, and various methods are currently being developed to overcome this problem, including oleosin-fusion technology, which allows extraction with oil bodies. In some cases, delivery of a biopharmaceutical product by direct ingestion of the modified plant potentially removes the need for purification. Such biopharmaceuticals and edible vaccines can be stored and distributed as seeds, tubers, or fruits, making immunization programs in developing countries cheaper and potentially easier to administer. Some of the most expensive biopharmaceuticals of restricted availability, such as glucocerebrosidase, could become much cheaper and more plentiful through production in transgenic plants. PMID:11062432

  18. [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.

  19. 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. PMID:22211569

  20. 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. PMID:18161495

  1. 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.

  2. [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. PMID:19722000

  3. 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. ...

  4. 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…

  5. 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...

  6. 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...

  7. Field tests of transgenic barley lines in North Dakota

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Testing transgenic barley lines for FHB in the greenhouse does not necessarily give the same results as field tests. The objective of this project was to test 18 transgenic lines in replicated trials in an inoculated FHB nursery. Several programs have developed barley lines expressing anti-fungal a...

  8. Public reactions and scientific responses to transgenic crops.

    PubMed

    Dale, P J

    1999-04-01

    There is currently intense debate in parts of Europe about the commercial production of transgenic food crops. Information from the press and lobbying groups has not encouraged an informed and balanced consideration of the issues. In marked contrast, there is widespread acceptance of transgenic food crops in North America.

  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. [Transgene complete silencing may associate with rearrangement of retroviral vector].

    PubMed

    Wang, Dan; Xiao, Lejia; Ma, Qingxin; Zhai, Fen; Ren, Sichong; Li, Changlong

    2011-04-01

    Transgene silencing is one of two major obstacles in both basic biomedical research for transgene and clinical practice of gene therapy. Based on the model of HT1080 cell clones, which transduced single copy of retroviral vector MGPN2, the mechanism of transgene silencing was explored in this investigation by a serial molecular techniques. In the HT1080 cell clone that absence of GFP protein synthesized, no significant aberration of epigenetic modification was detected, but the transcript size and the sequence changed that resulted in the reading frame shift. In addition to chromosomal position effect leading to transgene silencing, the transcript reading frame shift associated with retroviral vector rearrangements could induce complete silencing of transgene.

  11. Advancing environmental risk assessment for transgenic biofeedstock crops.

    PubMed

    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

  12. 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

  13. Advancing environmental risk assessment for transgenic biofeedstock crops.

    PubMed

    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.

  14. [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.

  15. 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. PMID:27565868

  16. 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

  17. Colorado potato beetles compensate for tomato cathepsin D inhibitor expressed in transgenic potato.

    PubMed

    Brunelle, France; Cloutier, Conrad; Michaud, Dominique

    2004-03-01

    We reported earlier the importance of digestive cathepsin D-like activity for initiating dietary protein hydrolysis in Colorado potato beetle, Leptinotarsa decemlineata Say [Brunelle et al. (1999) Arch. Insect Biochem. Physiol. 42:88-98]. We assessed here whether transgenic lines of potato (Solanum tuberosum L.) expressing a cathepsin D inhibitor (CDI) from tomato would show resistance to the beetle, or if the insect would compensate for the loss of cathepsin D activity after ingesting the recombinant inhibitor. Transgenic potato lines expressing tomato CDI were developed by Agrobacterium tumefaciens genetic transformation, and selected based on their relative amount of CDI. After confirming the absence of detectable visible effects of CDI on the plant's phenotype, diet assays with control and transgenic lines were carried out to assess the impact of the inhibitor on growth and development of the insect. Leaf consumption, relative growth rate, molting incidence, and digestive protease activity were monitored at 12-h intervals over 132 h for 3rd-instar larvae provided with transgenic potato foliage. Leaf consumption and relative growth rate were slightly reduced during the first 12 h for larvae fed CDI, but no significant differences were observed thereafter. In contrast, time for molting to the 4th larval stage was significantly longer for larvae fed modified plants, with developmental delays of approximately 10 h (0.5 day) compared to control larvae. Recombinant CDI also had an impact on the insect's digestive physiology, readily inducing overproduction of digestive proteases (rubiscases), followed by a gradual decrease of total and pepstatin-sensitive activity. Overall, these observations show the ability of Colorado potato beetle to compensate for the loss of cathepsin D activity by modulating its digestive protease complement in response to aspartate-type inhibitors in the diet. From a practical viewpoint, these data stress the importance of devising improved

  18. Cell cycle perturbation in the hepatocytes of HCV core transgenic mice following common bile duct ligation is associated with enhanced p21 expression.

    PubMed

    Chang, Ming-Ling; Chen, Tsung-Hsing; Chang, Ming-Yu; Yeh, Chau-Ting

    2009-03-01

    Hepatitis C virus (HCV) core protein has been reported to alter the cell cycle in vitro, but the data remain inconclusive, and in vitro experiments do not represent precisely events that occur in vivo, which may involve hepatic inflammation or regeneration. A group of double-transgenic mice carrying tetracycline transactivator (tTA) and HCV core that express conditionally the HCV core in the mature liver, and single-transgenic mice carrying only tTA were subjected to sham laparotomy, 43% partial hepatectomy, or common bile duct ligation. The cell cycle markers, including cyclin A, B1, D, E1, Mcm-2, phosphorylated histone 3 protein, Ki67, and p21Waf1/Cip1/Sdi1 (p21), were evaluated in liver samples obtained 3 days after the operation. No significant differences in the levels of any markers were observed between the double- and single-transgenic mice following sham laparotomy. Among the mice that underwent common bile duct ligation, the double-transgenic mice had lower levels of Ki67 (P = 0.0001), higher levels of cyclin D1 (P = 0.0001), and higher levels of p21 expression than the single-transgenic mice. Among those that underwent partial hepatectomy, the double-transgenic mice had higher p21 expression levels, but no significant differences in the levels of any other markers were observed between the double- and single-transgenic mice. It is concluded that the HCV core alters the hepatocyte cell cycle in addition to inducing G1 arrest in vivo after common bile duct ligation, and is associated with enhanced p21 expression. This may account at least partially for the strong association between HCV-related hepatocarcinogenesis and hepatic inflammation/ fibrosis.

  19. Comparative transcriptional and proteomic profiling of bread wheat cultivar and its derived transgenic line over-expressing a low molecular weight glutenin subunit gene in the endosperm

    Technology Transfer Automated Retrieval System (TEKTRAN)

    We have carried out a parallel transcriptional and proteomic comparison of seeds from a transformed bread wheat line that over-expresses a transgenic low molecular weight glutenin subunit gene relative to the corresponding non-transformed genotype. Proteomic analyses showed that, during seed develop...

  20. 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,...

  1. Cardiac phenotype induced by a dysfunctional α 1C transgene: a general problem for the transgenic approach.

    PubMed

    Asemu, Girma; Fishbein, Kenneth; Lao, Qi Zong; Ravindran, Arippa; Herbert, Ron; Canuto, Holly C; Spencer, Richard G; Soldatov, Nikolai M

    2011-01-01

    Based on stable integration of recombinant DNA into a host genome, transgenic technology has become an important genetic engineering methodology. An organism whose genetic characteristics have been altered by the insertion of foreign DNA is supposed to exhibit a new phenotype associated with the function of the transgene. However, successful insertion may not be sufficient to achieve specific modification of function. In this study we describe a strain of transgenic mouse, G7-882, generated by incorporation into the mouse genome of human CaV 1.2 α(1C) cDNA deprived of 3'-UTR to exclude transcription. We found that, in response to chronic infusion of isoproterenol, G7-882 develops dilated cardiomyopathy, a misleading "transgenic artifact" compatible with the expected function of the incorporated "correct" transgene. Specifically, using magnetic resonance imaging (MRI), we found that chronic β-adrenergic stimulation of G7-882 mice caused left ventricular hypertrophy and aggravated development of dilated cardiomyopathy, although no significant changes in the kinetics, density and voltage dependence of the calcium current were observed in G7-882 cardiomyocytes as compared to cells from wild type mice. This result illustrates the possibility that even when a functional transgene is expressed, an observed change in phenotype may be due to the artifact of "incidental incorporation" leading to misleading conclusions. To exclude this possibility and thus provide a robust tool for exploring biological function, the new transgenic phenotype must be replicated in several independently generated transgenic strains. PMID:21224729

  2. Cardiac phenotype induced by a dysfunctional α 1C transgene: a general problem for the transgenic approach.

    PubMed

    Asemu, Girma; Fishbein, Kenneth; Lao, Qi Zong; Ravindran, Arippa; Herbert, Ron; Canuto, Holly C; Spencer, Richard G; Soldatov, Nikolai M

    2011-01-01

    Based on stable integration of recombinant DNA into a host genome, transgenic technology has become an important genetic engineering methodology. An organism whose genetic characteristics have been altered by the insertion of foreign DNA is supposed to exhibit a new phenotype associated with the function of the transgene. However, successful insertion may not be sufficient to achieve specific modification of function. In this study we describe a strain of transgenic mouse, G7-882, generated by incorporation into the mouse genome of human CaV 1.2 α(1C) cDNA deprived of 3'-UTR to exclude transcription. We found that, in response to chronic infusion of isoproterenol, G7-882 develops dilated cardiomyopathy, a misleading "transgenic artifact" compatible with the expected function of the incorporated "correct" transgene. Specifically, using magnetic resonance imaging (MRI), we found that chronic β-adrenergic stimulation of G7-882 mice caused left ventricular hypertrophy and aggravated development of dilated cardiomyopathy, although no significant changes in the kinetics, density and voltage dependence of the calcium current were observed in G7-882 cardiomyocytes as compared to cells from wild type mice. This result illustrates the possibility that even when a functional transgene is expressed, an observed change in phenotype may be due to the artifact of "incidental incorporation" leading to misleading conclusions. To exclude this possibility and thus provide a robust tool for exploring biological function, the new transgenic phenotype must be replicated in several independently generated transgenic strains.

  3. Targeted transgenic expression of the mutation causing Hutchinson-Gilford progeria syndrome leads to proliferative and degenerative epidermal disease.

    PubMed

    Sagelius, Hanna; Rosengardten, Ylva; Hanif, Mubashir; Erdos, Michael R; Rozell, Björn; Collins, Francis S; Eriksson, Maria

    2008-04-01

    Hutchinson-Gilford progeria syndrome (HGPS) is a rare human genetic disorder characterized by striking progeroid features. Clinical findings in the skin include scleroderma, alopecia and loss of subcutaneous fat. HGPS is usually caused by a dominant-negative mutation in LMNA, a gene that encodes two major proteins of the inner nuclear lamina: lamin A and lamin C. We have generated tetracycline-inducible transgenic lines that carry a minigene of human LMNA under the control of a tet-operon. Two mouse lines were created: one carrying the wild-type sequence of LMNA and the other carrying the most common HGPS mutation. Targeted expression of the HGPS mutation in keratin-5-expressing tissues led to abnormalities in the skin and teeth, including fibrosis, loss of hypodermal adipocytes, structural defects in the hair follicles and sebaceous glands, and abnormal incisors. The severity of the defects was related to the level of expression of the transgene in different mouse lines. These transgenic mice appear to be good models for studies of the molecular mechanisms of skin abnormalities in HGPS and other related disorders.

  4. 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

  5. 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.

  6. 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

  7. Hmga2 promoter analysis in transgenic mice.

    PubMed

    Schiltz, John F; Rustighi, Alessandra; Tessari, Michela A; Liu, Jun; Braghetta, Paola; Sgarra, Riccardo; Stebel, Marco; Bressan, Giorgio M; Altruda, Fiorella; Giancotti, Vincenzo; Chada, Kiran; Manfioletti, Guidalberto

    2003-10-01

    HMGA2(2) belongs to the high mobility group A (HMGA) family of architectural transcription factors which participate in a wide variety of nuclear processes ranging from transcription to recombination, playing an important role in chromatin remodelling. HMGA2 is expressed during embryogenesis but not by adult somatic tissues, yet it becomes re-expressed following neoplastic transformation. A role in development is underscored by the finding that the inactivation of the Hmga2 gene is responsible for the murine pygmy phenotype. To elucidate mechanisms that control HMGA2 expression, we have previously cloned the gene and identified functional elements involved in its regulation. In this paper, transgenic mice were generated to define genomic regions involved in Hmga2 developmental and tissue-specific transcriptional regulation. A genomic region from -8.1 to -3.7kb upstream from the initiation site has been found to recapitulate most of the spatial and temporal endogenous Hmga2 gene expression. PMID:13679031

  8. Transcriptomic analyses of Hand2 transgenic embryos.

    PubMed

    Funato, Noriko; Kokubo, Hiroki; Saga, Yumiko

    2016-09-01

    In this article, we further provide the data generated for the previously published research article "Specification of jaw identity by the Hand2 transcription factor." To better understand the downstream genes of the basic helix-loop-helix transcription factor Hand2, we generated double-transgenic mice (Hand2 (NC) ) by intercrossing CAG-floxed CAT-Hand2 mice with Wnt1-Cre mice for conditional activation of Hand2 expression in the neural crest. Altered expression of Hand2 induces transformation of the upper jaw to the lower jaw in Hand2 (NC) mutant mice. This data article provides Tables detailing the differentially expressed genes between wild-type and Hand2 (NC) mutant embryos. The raw array data of our transcriptomes as generated using Affymetrix microarrays are available on the NCBI Gene Expression Omnibus (GEO) browser (Reference number GSE75805). PMID:27408813

  9. 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.

  10. 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.

  11. 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.

  12. 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.

  13. Partial rescue of insulin receptor-deficient mice by transgenic complementation with an activated insulin receptor in the liver.

    PubMed

    Baudry, Anne; Jackerott, Malene; Lamothe, Betty; Kozyrev, Sergey V; Leroux, Loïc; Durel, Béatrice; Saint-Just, Susan; Joshi, Rajiv L

    2002-10-16

    Insulin receptor (IR)-deficient mice develop severe diabetes mellitus, diabetic ketoacidosis (DKA) and liver steatosis and die within 1 week after birth. We examined in this work whether the metabolic phenotype of IR(-/-) mutants could be improved by transgenic complementation with IR selectively in the liver. We first generated transgenic mice expressing a human DNA complementary to RNA encoding a truncated constitutively activated form of IR (IRdelta) under the control of liver-specific phenylalanine hydroxylase (PAH) gene promoter. These mice presented more pronounced fasting hypoglycemia and showed slightly improved glucose tolerance as compared to controls. The transgenic mice were crossed with IR(+/-) mutants to generate IR(-/-) mice carrying the PAH-IRDelta transgene. Although such mutants developed glycosuria, DKA was delayed by more than 1 week and survival was prolonged to 8-20 days in approximately 10% of mice. In these partially rescued pups, serum glucose and triglyceride levels were lowered, hepatic glycogen stores were reconstituted and liver steatosis was absent as compared with pups which developed strong DKA and died earlier. Thus, lack of insulin action in the liver is responsible in large part for the metabolic disorders seen in IR(+/-) mice. This study should stimulate interest in therapeutic strategies aimed at improving hepatic function in diabetes.

  14. Transgenic expression of the endothelin-B receptor prevents congenital intestinal aganglionosis in a rat model of Hirschsprung disease.

    PubMed Central

    Gariepy, C E; Williams, S C; Richardson, J A; Hammer, R E; Yanagisawa, M

    1998-01-01

    The spotting lethal rat, a naturally occurring rodent model of Hirschsprung disease, carries a deletion in the endothelin-B receptor (EDNRB) gene that abrogates expression of functional EDNRB receptors. Rats homozygous for this mutation (sl) exhibit coat color spotting and congenital intestinal aganglionosis. These deficits result from failure of the neural crest-derived epidermal melanoblasts and enteric nervous system (ENS) precursors to completely colonize the skin and intestine, respectively. We demonstrate that during normal rat development, the EDNRB mRNA expression pattern is consistent with expression by ENS precursors throughout gut colonization. We used the human dopamine-beta-hydroxylase (DbetaH) promoter to direct transgenic expression of EDNRB to colonizing ENS precursors in the sl/sl rat. The DbetaH-EDNRB transgene compensates for deficient endogenous EDNRB in these rats and prevents the intestinal defect. The transgene has no effect on coat color spotting, indicating the critical time for EDNRB expression in enteric nervous system development begins after separation of the melanocyte lineage from the ENS lineage and their common precursor. The transgene dosage affects both the incidence and severity of the congenital intestinal defect, suggesting dosage-dependent events downstream of EDNRB activation in ENS development. PMID:9739043

  15. 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. PMID:26670088

  16. 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.

  17. 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.

  18. Gene flow from transgenic common beans expressing the bar gene.

    PubMed

    Faria, Josias C; Carneiro, Geraldo E S; Aragão, Francisco J L

    2010-01-01

    Gene flow is a common phenomenon even in self-pollinated plant species. With the advent of genetically modified plants this subject has become of the utmost importance due to the need for controlling the spread of transgenes. This study was conducted to determine the occurrence and intensity of outcrossing in transgenic common beans. In order to evaluate the outcross rates, four experiments were conducted in Santo Antonio de Goiás (GO, Brazil) and one in Londrina (PR, Brazil), using transgenic cultivars resistant to the herbicide glufosinate ammonium and their conventional counterparts as recipients of the transgene. Experiments with cv. Olathe Pinto and the transgenic line Olathe M1/4 were conducted in a completely randomized design with ten replications for three years in one location, whereas the experiments with cv. Pérola and the transgenic line Pérola M1/4 were conducted at two locations for one year, with the transgenic cultivar surrounded on all sides by the conventional counterpart. The outcross occurred at a negligible rate of 0.00741% in cv. Pérola, while none was observed (0.0%) in cv. Olathe Pinto. The frequency of gene flow was cultivar dependent and most of the observed outcross was within 2.5 m from the edge of the pollen source. Index terms: Phaseolus vulgaris, outcross, glufosinate ammonium. PMID:21865877

  19. Stability of the MON 810 transgene in maize.

    PubMed

    La Paz, Jose Luis; Pla, Maria; Papazova, Nina; Puigdomènech, Pere; Vicient, Carlos M

    2010-12-01

    We analysed the DNA variability of the transgene insert and its flanking regions in maize MON 810 commercial varieties. Southern analysis demonstrates that breeding, since the initial transformation event more than 10 years ago, has not resulted in any rearrangements. A detailed analysis on the DNA variability at the nucleotide level, using DNA mismatch endonuclease assays, showed the lack of polymorphisms in the transgene insert. We conclude that the mutation rate of the transgene is not significantly different from that observed in the maize endogenous genes. Six SNPs were observed in the 5'flanking region, corresponding to a Zeon1 retrotransposon long terminal repeat. All six SNPs are more than 500 bp upstream of the point of insertion of the transgene and do not affect the reliability of the established PCR-based transgene detection and quantification methods. The mutation rate of the flanking region is similar to that expected for a maize repetitive sequence. We detected low levels of cytosine methylation in leaves of different transgenic varieties, with no significant differences on comparing different transgenic varieties, and minor differences in cytosine methylation when comparing leaves at different developmental stages. There was also a reduction in cryIAb mRNA accumulation during leaf development. PMID:20936423

  20. Production of transgenic miniature pigs by pronuclear microinjection.

    PubMed

    Uchida, M; Shimatsu, Y; Onoe, K; Matsuyama, N; Niki, R; Ikeda, J E; Imai, H

    2001-12-01

    Miniature pig is an attractive animal for a wide range of research fields, such as medicine and pharmacology, because of its small size, the possibility of breeding it under minimum environmental controls and the physiology that is potentially similar to that of human. Although transgenic technology is useful for the analysis of gene function and for the development of model animals for various diseases, there have not yet been any reports on producing transgenic miniature pig. This study is the first successful report concerning the production of transgenic miniature pig by pronuclear microinjection. The huntingtin gene cloned from miniature pig, which is a homologue of candidate gene for Huntington's disease, connected with rat neuron-specific enolase promoter region, was injected into a pronucleus of fertilized eggs with micromanipulator. The eggs were transferred into the oviduct of recipient miniature pigs, whose estrus cycles were previously synchronized with a progesterone analogue. A total of 402 injected eggs from 171 donors were transferred to 23 synchronized recipients. Sixteen of them maintained pregnancy and delivered 65 young, and one resulted in abortion. Five of the 68 offspring (three of which were aborted) were determined to have transgene by PCR and Southern analysis. The overall rate of transgenic production was 1.24% (transgenic/injected eggs). This study provides the first success and useful information regarding production of transgenic miniature pig for biomedical research.

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

    PubMed

    Huber, Reinhard C; Remuge, Liliana; Carlisle, Ailsa; Lillico, Simon; Sandøe, Peter; Sørensen, Dorte B; Whitelaw, C Bruce A; Olsson, I Anna S

    2012-08-01

    Since large animal transgenesis has been successfully attempted for the first time about 25 years ago, the technology has been applied in various lines of transgenic pigs. Nevertheless one of the concerns with the technology--animal welfare--has not been approached through systematic assessment and statements regarding the welfare of transgenic pigs have been based on anecdotal observations during early stages of transgenic programs. The main aim of the present study was therefore to perform an extensive welfare assessment comparing heterozygous transgenic animals expressing GFP with wildtype animals along various stages of post natal development. The protocol used covered reproductory performance and behaviour in GFP and wildtype sows and general health and development, social behaviour, exploratory behaviour and emotionality in GFP and wildtype littermates from birth until an age of roughly 4 months. The absence of significant differences between GFP and wildtype animals in the parameters observed suggests that the transgenic animals in question are unlikely to suffer from deleterious effects of transgene expression on their welfare and thus support existing anecdotal observations of pigs expressing GFP as healthy. Although the results are not surprising in the light of previous experience, they give a more solid fundament to the evaluation of GFP expression as being relatively non-invasive in pigs. The present study may furthermore serve as starting point for researchers aiming at a systematic characterization of welfare relevant effects in the line of transgenic pigs they are working with.

  2. Genetic load and transgenic mitigating genes in transgenic Brassica rapa (field mustard) × Brassica napus (oilseed rape) hybrid populations

    PubMed Central

    2009-01-01

    Background One theoretical explanation for the relatively poor performance of Brassica rapa (weed) × Brassica napus (crop) transgenic hybrids suggests that hybridization imparts a negative genetic load. Consequently, in hybrids genetic load could overshadow any benefits of fitness enhancing transgenes and become the limiting factor in transgenic hybrid persistence. Two types of genetic load were analyzed in this study: random/linkage-derived genetic load, and directly incorporated genetic load using a transgenic mitigation (TM) strategy. In order to measure the effects of random genetic load, hybrid productivity (seed yield and biomass) was correlated with crop- and weed-specific AFLP genomic markers. This portion of the study was designed to answer whether or not weed × transgenic crop hybrids possessing more crop genes were less competitive than hybrids containing fewer crop genes. The effects of directly incorporated genetic load (TM) were analyzed through transgene persistence data. TM strategies are proposed to decrease transgene persistence if gene flow and subsequent transgene introgression to a wild host were to occur. Results In the absence of interspecific competition, transgenic weed × crop hybrids benefited from having more crop-specific alleles. There was a positive correlation between performance and number of B. napus crop-specific AFLP markers [seed yield vs. marker number (r = 0.54, P = 0.0003) and vegetative dry biomass vs. marker number (r = 0.44, P = 0.005)]. However under interspecific competition with wheat or more weed-like conditions (i.e. representing a situation where hybrid plants emerge as volunteer weeds in subsequent cropping systems), there was a positive correlation between the number of B. rapa weed-specific AFLP markers and seed yield (r = 0.70, P = 0.0001), although no such correlation was detected for vegetative biomass. When genetic load was directly incorporated into the hybrid genome, by inserting a fitness

  3. 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.

  4. [Effects of phytase transgenic corn planting on soil nematode community].

    PubMed

    Zhao, Zong-Chao; Su, Ying; Mou, Wen-Ya; Liu, Man-Qiang; Chen, Xiao-Yun; Chen, Fa-Jun

    2014-04-01

    A healthy soil ecosystem is essential for nutrient cycling and energy conversion, and the impact of exogenous genes from genetically modified crops had aroused wide concerns. Phytase transgenic corn (i. e., the inbred line BVLA430101) was issued a bio-safety certificate on 27 September 2009 in China, which could improve the efficiency of feed utilization, reduce environmental pollution caused by animal manure. In this study, the abundance of trophic groups, community structure and ecological indices of soil nematodes were studied over the growing cycle of phytase transgenic corn (ab. transgenic corn) and control conventional parental corn (ab. control corn) in the field. Totally 29 and 26 nematode genera were isolated from transgenic corn and control corn fields, respectively. The abundances of bacterivores and omnivores-predators, the total number of soil nematodes, and the Shannon index (H) were significantly greater under transgenic corn than under control corn, while the opposite trend was found for the relative abundance of herbivores and the maturity index (Sigma MI) of soil nematodes. Repeated-measures analysis of variance (ANOVA) did not detect any significant effects of transgenic corn on the composition and abundance of nematode trophic groups and ecological indices of soil nematodes. Furthermore, the Student-T test showed that the abundances of bacterivores and omnivores-predators and the total number of soil nematodes during the milk-ripe stage were significant higher in the transgenic corn field than in the control corn field. The effects of transgenic corn planting on soil nematodes might be related to the increase in the nitrogen content of field soil under transgenic corn compared to control corn.

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

    PubMed

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

    1997-09-01

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

  6. [Effects of phytase transgenic corn planting on soil nematode community].

    PubMed

    Zhao, Zong-Chao; Su, Ying; Mou, Wen-Ya; Liu, Man-Qiang; Chen, Xiao-Yun; Chen, Fa-Jun

    2014-04-01

    A healthy soil ecosystem is essential for nutrient cycling and energy conversion, and the impact of exogenous genes from genetically modified crops had aroused wide concerns. Phytase transgenic corn (i. e., the inbred line BVLA430101) was issued a bio-safety certificate on 27 September 2009 in China, which could improve the efficiency of feed utilization, reduce environmental pollution caused by animal manure. In this study, the abundance of trophic groups, community structure and ecological indices of soil nematodes were studied over the growing cycle of phytase transgenic corn (ab. transgenic corn) and control conventional parental corn (ab. control corn) in the field. Totally 29 and 26 nematode genera were isolated from transgenic corn and control corn fields, respectively. The abundances of bacterivores and omnivores-predators, the total number of soil nematodes, and the Shannon index (H) were significantly greater under transgenic corn than under control corn, while the opposite trend was found for the relative abundance of herbivores and the maturity index (Sigma MI) of soil nematodes. Repeated-measures analysis of variance (ANOVA) did not detect any significant effects of transgenic corn on the composition and abundance of nematode trophic groups and ecological indices of soil nematodes. Furthermore, the Student-T test showed that the abundances of bacterivores and omnivores-predators and the total number of soil nematodes during the milk-ripe stage were significant higher in the transgenic corn field than in the control corn field. The effects of transgenic corn planting on soil nematodes might be related to the increase in the nitrogen content of field soil under transgenic corn compared to control corn. PMID:25011306

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

    PubMed

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

    2012-01-01

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

  8. Generation and Characterization of Human Heme Oxygenase-1 Transgenic Pigs

    PubMed Central

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

    2012-01-01

    Xenotransplantation using transgenic pigs as an organ source is a promising strategy to overcome shortage of human organ for transplantation. Various genetic modifications have been tried to ameliorate xenograft rejection. In the present study we assessed effect of transgenic expression of human heme oxygenase-1 (hHO-1), an inducible protein capable of cytoprotection by scavenging reactive oxygen species and preventing apoptosis caused by cellular stress during inflammatory processes, in neonatal porcine islet-like cluster cells (NPCCs). Transduction of NPCCs with adenovirus containing hHO-1 gene significantly reduced apoptosis compared with the GFP-expressing adenovirus control after treatment with either hydrogen peroxide or hTNF-α and cycloheximide. These protective effects were diminished by co-treatment of hHO-1 antagonist, Zinc protoporphyrin IX. We also generated transgenic pigs expressing hHO-1 and analyzed expression and function of the transgene. Human HO-1 was expressed in most tissues, including the heart, kidney, lung, pancreas, spleen and skin, however, expression levels and patterns of the hHO-1 gene are not consistent in each organ. We isolate fibroblast from transgenic pigs to analyze protective effect of the hHO-1. As expected, fibroblasts derived from the hHO-1 transgenic pigs were significantly resistant to both hydrogen peroxide damage and hTNF-α and cycloheximide-mediated apoptosis when compared with wild-type fibroblasts. Furthermore, induction of RANTES in response to hTNF-α or LPS was significantly decreased in fibroblasts obtained from the hHO-1 transgenic pigs. These findings suggest that transgenic expression of hHO-1 can protect xenografts when exposed to oxidative stresses, especially from ischemia/reperfusion injury, and/or acute rejection mediated by cytokines. Accordingly, hHO-1 could be an important candidate molecule in a multi-transgenic pig strategy for xenotransplantation. PMID:23071605

  9. 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. PMID:26160901

  10. 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.

  11. Endogenous allergen upregulation: transgenic vs. traditionally bred crops.

    PubMed

    Herman, Rod A; Ladics, Gregory S

    2011-10-01

    The safety assessment for transgenic food crops currently includes an evaluation of the endogenous allergy potential (via serum IgE screening) when the non-transgenic counterpart is a commonly allergenic food. The value of this analysis in the safety assessment of transgenic crops, especially with reference to recent requests to quantify individual allergen concentrations in raw commodities, is examined. We conclude that the likelihood of upregulating an endogenous allergen due to transgenesis is no greater than from traditional breeding which has a history of safety and is largely unregulated. The potential consequences of upregulating an endogenous allergen are also unclear.

  12. Transgenic plant-derived pharmaceuticals - the practical approach?

    PubMed

    Yano, Akira; Takekoshi, Masataka

    2004-10-01

    Production of biopharmaceuticals in transgenic plants would involve the creation of a new industry. Those transgenic plants, including staple food crops, could provide many benefits to people all over the world. However, the new industry might require a strict regulation system. It is probable that such a strict system would not be acceptable to Japan or to most developing countries. Many countries should use non-food crops for production of biopharmaceuticals and take on more simple systems. The new industry must develop strategies for promoting the benefits of transgenic plant-derived biopharmaceuticals on both the domestic and worldwide scales.

  13. 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

  14. The effect of Bt-transgene introgression on plant growth and reproduction in wild Brassica juncea.

    PubMed

    Liu, Yong-Bo; Darmency, Henry; Stewart, C Neal; Wei, Wei; Tang, Zhi-Xi; Ma, Ke-Ping

    2015-06-01

    This study aims to investigate the relative plant growth and reproduction of insect-resistant and susceptible plants following the introgression of an insect-resistance Bt-transgene from Brassica napus, oilseed rape, to wild Brassica juncea. The second backcrossed generation (BC2) from a single backcross family was grown in pure and mixed stands of Bt-transgenic and non-transgenic siblings under two insect treatments. Various proportions of Bt-transgenic plants were employed in mixed stands to study the interaction between resistant and susceptible plants. In the pure stands, Bt-transgenic BC2 plants performed better than non-transgenic plants with or without insect treatments. In mixed stands, Bt-transgenic BC2 plants produced fewer seeds than their non-Bt counterparts at low proportions of Bt-transgenic BC2 plants in the absence of insects. Reproductive allocation of non-transgenic plants marginally increased with increasing proportions of Bt-transgenic plants under herbivore pressure, which resulted in increased total biomass and seed production per stand. The results showed that the growth of non-transgenic plants was protected by Bt-transgenic plants under herbivore pressure. The Bt-transgene might not be advantageous in mixed stands of backcrossed hybrids; thus transgene introgression would not be facilitated when herbivorous insects are not present. However, a relatively large initial population of Bt-transgenic plants might result in transgene persistence when target herbivores are present. PMID:25487040

  15. The effect of Bt-transgene introgression on plant growth and reproduction in wild Brassica juncea.

    PubMed

    Liu, Yong-Bo; Darmency, Henry; Stewart, C Neal; Wei, Wei; Tang, Zhi-Xi; Ma, Ke-Ping

    2015-06-01

    This study aims to investigate the relative plant growth and reproduction of insect-resistant and susceptible plants following the introgression of an insect-resistance Bt-transgene from Brassica napus, oilseed rape, to wild Brassica juncea. The second backcrossed generation (BC2) from a single backcross family was grown in pure and mixed stands of Bt-transgenic and non-transgenic siblings under two insect treatments. Various proportions of Bt-transgenic plants were employed in mixed stands to study the interaction between resistant and susceptible plants. In the pure stands, Bt-transgenic BC2 plants performed better than non-transgenic plants with or without insect treatments. In mixed stands, Bt-transgenic BC2 plants produced fewer seeds than their non-Bt counterparts at low proportions of Bt-transgenic BC2 plants in the absence of insects. Reproductive allocation of non-transgenic plants marginally increased with increasing proportions of Bt-transgenic plants under herbivore pressure, which resulted in increased total biomass and seed production per stand. The results showed that the growth of non-transgenic plants was protected by Bt-transgenic plants under herbivore pressure. The Bt-transgene might not be advantageous in mixed stands of backcrossed hybrids; thus transgene introgression would not be facilitated when herbivorous insects are not present. However, a relatively large initial population of Bt-transgenic plants might result in transgene persistence when target herbivores are present.

  16. Long-term toxicity study on transgenic rice with Cry1Ac and sck genes.

    PubMed

    Zhang, Min; Zhuo, Qin; Tian, Yuan; Piao, Jianhua; Yang, Xiaoguang

    2014-01-01

    In the present work, we evaluated the chronic effects of the transgenic insect-resistant rice carrying Cry1Ac and sck genes on Sprague-Dawley (SD) rats through a 78-week feeding study. Based on the gender and weight, 180 SD rats were randomly and evenly assigned into three groups. GM rice and non-GM rice were separately formulated into diets at high levels. AIN-93 diet was used as a nutritional control. Body weight, food consumption, hematology and serum chemistry were monitored regularly. Rats were sacrificed for organ weight measurement and pathological examination at 52 weeks and 78 weeks. Body weight, food consumption, mortality rates, tumor incidences and pathological findings showed no significant difference among the three groups. Although certain differences in some hematology, serum chemistry parameters and relative organ weights were observed between GM rice group and control groups, they were not considered as treatment-related. Taken together, long-term intake of transgenic rice carrying Cry1Ac and sck genes at a high level exerts no unintended adverse effects on rats.

  17. The role of working memory in carrying and borrowing.

    PubMed

    Imbo, Ineke; Vandierendonck, André; Vergauwe, Evie

    2007-07-01

    The present study analyzed the role of phonological and executive components of working memory in the borrow operation in complex subtractions (Experiments 1 and 2) and in the carry operation in complex multiplications (Experiments 3 and 4). The number of carry and borrow operations as well as the value of the carry were manipulated. Results indicated that both the number of carry/borrow operations and the value of the carry increased problem difficulty, resulting in higher reliance on phonological and executive working-memory components. Present results are compared with those obtained for the carry operation in complex addition and are further discussed in the broader framework of working-memory functions.

  18. Functional proteome of macrophage carried nanoformulated antiretroviral therapy demonstrates enhanced particle carrying capacity.

    PubMed

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

    2013-05-01

    Our laboratory developed long-acting nanoformulations of antiretroviral therapy (nanoART) to improve drug compliance, reduce toxicities, and facilitate access of drug to viral reservoirs. These all function to inevitably 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

  19. The Presence of a Chromatin Boundary Appears to Shield a Transgene in Tobacco from RNA Silencing

    PubMed Central

    Mlynárová, Ludmila; Hricová, Andrea; Loonen, Annelies; Nap, Jan-Peter

    2003-01-01

    We present isogenic transgenic tobacco lines that carry at a given chromosomal position a β-glucuronidase (GUS) reporter gene either with or without the presence of the matrix-associated region known as the chicken lysozyme A element. Plants were generated with the Cre-lox site–specific recombination system using heterospecific lox sites. Analysis of GUS gene expression in plant populations demonstrates that the presence of the A element can shield against RNA silencing of the GUS gene. Protection was observed in two of three independent tobacco transformants. Plants carrying an A element 5′ of the GUS gene always had stable GUS activity, but upon removal of this A element, the GUS gene became silenced over time in two lines, notably when homozygous. PMID:12953121

  20. Generation of Chlamydomonas strains that efficiently express nuclear transgenes.

    PubMed

    Neupert, Juliane; Karcher, Daniel; Bock, Ralph

    2009-03-01

    The unicellular green alga Chlamydomonas reinhardtii is both an invaluable model organism for plant biology and an attractive biotechnological production system. Despite the availability of efficient methods for introduction of foreign genes into the nuclear genome of the alga, transgene expression levels are usually very poor. This is a serious limitation that has severely hampered both post-genomics research in Chlamydomonas and use of the alga in molecular farming. Here we report a solution to this problem. We have designed a genetic screen that facilitates isolation of algal strains that efficiently express introduced transgenes. The levels of accumulation of foreign protein in our expression strains are almost uniformly high in all transgenic clones and are little influenced by position effects. The possibility of expressing transgenes to high levels will greatly facilitate post-genomics research in Chlamydomonas, and will also boost exploitation of the alga as an inexpensive production host for biopharmaceuticals and other valuable compounds.

  1. [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.

  2. Pronuclear Microinjection and Oviduct Transfer Procedures for Transgenic Mouse Production

    PubMed Central

    Liu, Chengyu; Xie, Wen; Gui, Changyun; Du, Yubin

    2013-01-01

    Transgenic mouse technology is a powerful method for studying gene function and creating animal models of human diseases. Currently, the most widely used method for generating transgenic mice is the pronuclear microinjection method. In this method, a transgenic DNA construct is physically microinjected into the pronucleus of a fertilized egg. The injected embryos are subsequently transferred into the oviducts of pseudopregnant surrogate mothers. A portion of the mice born to these surrogate mothers will harbor the injected foreign gene in their genomes. These procedures are technically challenging for most biomedical researchers. Inappropriate experimental procedures or suboptimal equipment setup can substantially reduce the efficiency of transgenic mouse production. In this chapter, we describe in detail our microinjection setup as well as our standard microinjection and oviduct transfer procedures. PMID:23912989

  3. [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. PMID:25757330

  4. Pronuclear microinjection and oviduct transfer procedures for transgenic mouse production.

    PubMed

    Liu, Chengyu; Xie, Wen; Gui, Changyun; Du, Yubin

    2013-01-01

    Transgenic mouse technology is a powerful method for studying gene function and creating animal models of human diseases. Currently, the most widely used method for generating transgenic mice is the pronuclear microinjection method. In this method, a transgenic DNA construct is physically microinjected into the pronucleus of a fertilized egg. The injected embryos are subsequently transferred into the oviducts of pseudopregnant surrogate mothers. A portion of the mice born to these surrogate mothers will harbor the injected foreign gene in their genomes. These procedures are technically challenging for most biomedical researchers. Inappropriate experimental procedures or suboptimal equipment setup can substantially reduce the efficiency of transgenic mouse production. In this chapter, we describe in detail our microinjection setup as well as our standard microinjection and oviduct transfer procedures. PMID:23912989

  5. 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 ...

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

    PubMed

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

    2014-04-25

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

  7. Microinjection of A. aegypti embryos to obtain transgenic mosquitoes.

    PubMed

    Jasinskiene, Nijole; Juhn, Jennifer; James, Anthony A

    2007-01-01

    In this video, Nijole Jasinskiene demonstrates the methodology employed to generate transgenic Aedes aegypti mosquitoes, which are vectors for dengue fever. The techniques for correctly preparing microinjection needles, desiccating embryos, and performing microinjection are demonstrated.

  8. Designer proton-channel transgenic algae for photobiological hydrogen production

    DOEpatents

    Lee, James Weifu

    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.

  9. 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...

  10. GhDRIN1, a novel drought-induced gene of upland cotton (Gossypium hirsutum L.) confers abiotic and biotic stress tolerance in transgenic tobacco.

    PubMed

    Dhandapani, Gurusamy; Lakshmi Prabha, Azhagiyamanavalan; Kanakachari, Mogilicherla; Phanindra, Mullapudi Lakshmi Venkata; Prabhakaran, Narayanasamy; Gothandapani, Sellamuthu; Padmalatha, Kethireddy Venkata; Solanke, Amolkumar U; Kumar, Polumetla Ananda

    2015-04-01

    A novel stress tolerance cDNA fragment encoding GhDRIN1 protein was identified and its regulation was studied in cotton boll tissues and seedlings subjected to various biotic and abiotic stresses. Phylogenetic and conserved domain prediction indicated that GhDRIN1 was annotated with a hypothetical protein of unknown function. Subcellular localization showed that GhDRIN1 is localized in the chloroplasts. The promoter sequence was isolated and subjected to in silico study. Various cis-acting elements responsive to biotic and abiotic stresses and hormones were found. Transgenic tobacco seedlings exhibited better growth on amended MS medium and showed minimal leaf damage in insect bioassays carried out with Helicoverpa armigera larvae. Transgenic tobacco showed better tolerance to water-deficit and fast recovered upon rewatering. Present work demonstrated that GhDRIN1, a novel stress tolerance gene of cotton, positively regulates the response to biotic and abiotic stresses in transgenic tobacco. PMID:25413882

  11. GhDRIN1, a novel drought-induced gene of upland cotton (Gossypium hirsutum L.) confers abiotic and biotic stress tolerance in transgenic tobacco.

    PubMed

    Dhandapani, Gurusamy; Lakshmi Prabha, Azhagiyamanavalan; Kanakachari, Mogilicherla; Phanindra, Mullapudi Lakshmi Venkata; Prabhakaran, Narayanasamy; Gothandapani, Sellamuthu; Padmalatha, Kethireddy Venkata; Solanke, Amolkumar U; Kumar, Polumetla Ananda

    2015-04-01

    A novel stress tolerance cDNA fragment encoding GhDRIN1 protein was identified and its regulation was studied in cotton boll tissues and seedlings subjected to various biotic and abiotic stresses. Phylogenetic and conserved domain prediction indicated that GhDRIN1 was annotated with a hypothetical protein of unknown function. Subcellular localization showed that GhDRIN1 is localized in the chloroplasts. The promoter sequence was isolated and subjected to in silico study. Various cis-acting elements responsive to biotic and abiotic stresses and hormones were found. Transgenic tobacco seedlings exhibited better growth on amended MS medium and showed minimal leaf damage in insect bioassays carried out with Helicoverpa armigera larvae. Transgenic tobacco showed better tolerance to water-deficit and fast recovered upon rewatering. Present work demonstrated that GhDRIN1, a novel stress tolerance gene of cotton, positively regulates the response to biotic and abiotic stresses in transgenic tobacco.

  12. Transgenic mouse model of malignant skin melanoma.

    PubMed Central

    Mintz, B; Silvers, W K

    1993-01-01

    Tyr-SV40E transgenic mice are specifically susceptible to melanoma due to expression of the oncogene in pigment cells. Mice of the more susceptible lines die young of early-onset eye melanomas, when skin melanomas are still infrequent and benign. To surmount this obstacle, skin from donors of two high-susceptibility lines was grafted to Tyr-SV40E hosts of a low-susceptibility line of the same inbred strain, thereby enabling the skin to outlive the donors and continue to grow in immunocompetent but tolerant hosts. Unexpectedly, donor pigment cells in all the grafts soon selectively proliferated close to areas of greatest wound healing, forming a dense black tracery, especially at the outer rim of the grafts. These lesions slowly grew radially within the grafts, producing irregular greyish patches. Local vertical thickenings then appeared and developed into small melanomas, which soon ulcerated through the epidermis. The tumors rapidly enlarged and became deeply invasive. Discrete black nevi also arose, with many becoming larger and distinctly blue, but those not near areas of pronounced wound healing did not progress to malignancy. In this first series, malignant melanoma resulted in all the grafts from the more susceptible of two donor lines and in some grafts from the other line. Distant metastases occurred in some cases from each line. Most tumors were hypomelanotic and heterogeneous, with lobes or areas differing in melanization. The results strongly suggest that growth factors and cytokines--known to be produced in wound repair--are triggering the growth and malignant conversion of these genetically susceptible melanocytes and that in the graft situation we are merely witnessing a caricature--a usefully exaggerated manifestation of the true events underlying the genesis of melanomas. The striking resemblance to the human malignancy, the genetic uniformity and different susceptibilities of the transgenic lines, and the experimental possibilities in the grafted

  13. Mutations in the coat protein gene of plum pox virus suppress particle assembly, heterologous encapsidation and complementation in transgenic plants of Nicotiana benthamiana.

    PubMed

    Varrelmann, M; Maiss, E

    2000-03-01

    Two different motifs in the coat protein (CP) of Plum pox virus (PPV) (R(3015)Q(3016), D(3059)) were mutated by replacing the respective amino acids with others possessing different chemical properties. The mutated CP genes were introduced into an infectious full-length clone of PPV (p35PPV-NAT) to investigate their influence on systemic infection of transgenic wild-type PPV CP-expressing and non-transgenic plants of Nicotiana benthamiana. All mutants failed to establish systemic infections in non-transgenic N. benthamiana plants, but were complemented by intact CP in transgenic plants. Moreover, the CP-RQ-D mutant (carrying mutations in both the RQ and D motifs) was introduced into p35PPV-NAT engineered to express beta-glucuronidase (GUS) for direct observation of systemic movement and particle assembly in N. benthamiana leaves. GUS-staining revealed that the CP mutant (RQ-D) was restricted to initially infected cells without forming virions. Systemic movement and particle assembly were restored in CP-transgenic N. benthamiana plants. Finally, transgenic N. benthamiana plants were generated that expressed each of the three mutated CP genes. Homozygous T(2) lines were selected and tested for resistance to PPV. Immunogold labelling and electron microscopy revealed that heterologous encapsidation with challenging Chilli veinal mottle virus and Potato virus Y was suppressed in these lines. In addition, assembly mutants did not complement CP-defective p35PPV-NAT. The possible use of modified viral CP genes for the production of virus-resistant transgenic plants, thereby reducing the putative risks of heterologous encapsidation and complementation, is discussed. PMID:10675394

  14. 46 CFR 169.213 - Permit to carry excursion party.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ..., the number of persons the vessel may carry, the crew required, and additional lifesaving or safety... permit to carry an excursion party must be in full compliance with the terms of its certificate...

  15. 46 CFR 176.204 - Permit to carry excursion party.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... may carry, the crew required, any additional lifesaving or safety equipment required, the route for... vessel operating under a permit to carry an excursion party must be in full compliance with the terms...

  16. Wheat chloroplast targeted sHSP26 promoter confers heat and abiotic stress inducible expression in transgenic Arabidopsis Plants.

    PubMed

    Khurana, Neetika; Chauhan, Harsh; Khurana, Paramjit

    2013-01-01

    The small heat shock proteins (sHSPs) have been found to play a critical role in physiological stress conditions in protecting proteins from irreversible aggregation. To characterize the hloroplast targeted sHSP26 promoter in detail, deletion analysis of the promoter is carried out and analysed via transgenics in Arabidopsis. In the present study, complete assessment of the importance of CCAAT-box elements along with Heat shock elements (HSEs) in the promoter of sHSP26 was performed. Moreover, the importance of 5' untranslated region (UTR) has also been established in the promoter via Arabidopsis transgenics. An intense GUS expression was observed after heat stress in the transgenics harbouring a full-length promoter, confirming the heat-stress inducibility of the promoter. Transgenic plants without UTR showed reduced GUS expression when compared to transgenic plants with UTR as was confirmed at the RNA and protein levels by qRT-PCR and GUS histochemical assays, thus suggesting the possible involvement of some regulatory elements present in the UTR in heat-stress inducibility of the promoter. Promoter activity was also checked under different abiotic stresses and revealed differential expression in different deletion constructs. Promoter analysis based on histochemical assay, real-time qPCR and fluorimetric analysis revealed that HSEs alone could not transcribe GUS gene significantly in sHSP26 promoter and CCAAT box elements contribute synergistically to the transcription. Our results also provide insight into the importance of 5`UTR of sHsp26 promoter thus emphasizing the probable role of imperfect CCAAT-box element or some novel cis-element with respect to heat stress. PMID:23349883

  17. Analysis of tomato polygalacturonase expression in transgenic tobacco.

    PubMed Central

    Osteryoung, K W; Toenjes, K; Hall, B; Winkler, V; Bennett, A B

    1990-01-01

    Tomato polygalacturonase is a cell wall enzyme secreted in large amounts during tomato fruit ripening. Polygalacturonase is synthesized as a glycoprotein precursor that undergoes numerous cotranslational and post-translational processing steps during its maturation, yielding three isozymes in tomato fruit, PG1, PG2A, and PG2B. To investigate the physiological roles of the three isozymes and the functional significance of the polygalacturonase processing domains in its intracellular transport and activity, we have examined polygalacturonase expression in transgenic tobacco plants. A full-length polygalacturonase cDNA was placed under control of the cauliflower mosaic virus 35S promoter and introduced into tobacco by way of Agrobacterium-mediated transformation. Analysis of transgenic tobacco plants indicated that (1) immunologically detectable polygalacturonase can be extracted from leaves, roots, and stems of transgenic tobacco plants; (2) only PG2A and PG2B were detectable in transgenic tobacco; (3) the polygalacturonase isozymes present in transgenic tobacco were electrophoretically indistinguishable from the tomato isozymes; (4) the N-terminal sequence, degree of N-linked glycosylation, and extent of oligosaccharide processing were similar in polygalacturonase from transgenic tobacco and tomato; (5) polygalacturonase was properly localized in cell walls of transgenic tissue; (6) the protein was enzymatically active in vitro; however, (7) accumulation of PG2A and PG2B in cell walls of transgenic tobacco did not result in pectin degradation in vivo. These results indicated that tomato polygalacturonase was properly processed and transported to the cell wall of tobacco. However, accumulation of the two polygalacturonase isozymes expressed in this heterologous host was insufficient to promote polyuronide degradation in tobacco leaf tissue. PMID:2152163

  18. 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. .

  19. 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.

  20. 30 CFR 57.16014 - Operator-carrying overhead cranes.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 30 Mineral Resources 1 2010-07-01 2010-07-01 false Operator-carrying overhead cranes. 57.16014 Section 57.16014 Mineral Resources MINE SAFETY AND HEALTH ADMINISTRATION, DEPARTMENT OF LABOR METAL AND... Storage and Handling § 57.16014 Operator-carrying overhead cranes. Operator-carrying overhead cranes...

  1. 30 CFR 56.16014 - Operator-carrying overhead cranes.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 30 Mineral Resources 1 2010-07-01 2010-07-01 false Operator-carrying overhead cranes. 56.16014 Section 56.16014 Mineral Resources MINE SAFETY AND HEALTH ADMINISTRATION, DEPARTMENT OF LABOR METAL AND... Storage and Handling § 56.16014 Operator-carrying overhead cranes. Operator-carrying overhead cranes...

  2. 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...

  3. 30 CFR 57.16014 - Operator-carrying overhead cranes.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 30 Mineral Resources 1 2014-07-01 2014-07-01 false Operator-carrying overhead cranes. 57.16014 Section 57.16014 Mineral Resources MINE SAFETY AND HEALTH ADMINISTRATION, DEPARTMENT OF LABOR METAL AND... Storage and Handling § 57.16014 Operator-carrying overhead cranes. Operator-carrying overhead cranes...

  4. 30 CFR 56.16014 - Operator-carrying overhead cranes.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 30 Mineral Resources 1 2014-07-01 2014-07-01 false Operator-carrying overhead cranes. 56.16014 Section 56.16014 Mineral Resources MINE SAFETY AND HEALTH ADMINISTRATION, DEPARTMENT OF LABOR METAL AND... Storage and Handling § 56.16014 Operator-carrying overhead cranes. Operator-carrying overhead cranes...

  5. 30 CFR 57.16014 - Operator-carrying overhead cranes.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 30 Mineral Resources 1 2013-07-01 2013-07-01 false Operator-carrying overhead cranes. 57.16014 Section 57.16014 Mineral Resources MINE SAFETY AND HEALTH ADMINISTRATION, DEPARTMENT OF LABOR METAL AND... Storage and Handling § 57.16014 Operator-carrying overhead cranes. Operator-carrying overhead cranes...

  6. 30 CFR 56.16014 - Operator-carrying overhead cranes.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 30 Mineral Resources 1 2013-07-01 2013-07-01 false Operator-carrying overhead cranes. 56.16014 Section 56.16014 Mineral Resources MINE SAFETY AND HEALTH ADMINISTRATION, DEPARTMENT OF LABOR METAL AND... Storage and Handling § 56.16014 Operator-carrying overhead cranes. Operator-carrying overhead cranes...

  7. 30 CFR 57.16014 - Operator-carrying overhead cranes.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 30 Mineral Resources 1 2012-07-01 2012-07-01 false Operator-carrying overhead cranes. 57.16014 Section 57.16014 Mineral Resources MINE SAFETY AND HEALTH ADMINISTRATION, DEPARTMENT OF LABOR METAL AND... Storage and Handling § 57.16014 Operator-carrying overhead cranes. Operator-carrying overhead cranes...

  8. 30 CFR 56.16014 - Operator-carrying overhead cranes.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 30 Mineral Resources 1 2012-07-01 2012-07-01 false Operator-carrying overhead cranes. 56.16014 Section 56.16014 Mineral Resources MINE SAFETY AND HEALTH ADMINISTRATION, DEPARTMENT OF LABOR METAL AND... Storage and Handling § 56.16014 Operator-carrying overhead cranes. Operator-carrying overhead cranes...

  9. 30 CFR 56.16014 - Operator-carrying overhead cranes.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 30 Mineral Resources 1 2011-07-01 2011-07-01 false Operator-carrying overhead cranes. 56.16014 Section 56.16014 Mineral Resources MINE SAFETY AND HEALTH ADMINISTRATION, DEPARTMENT OF LABOR METAL AND... Storage and Handling § 56.16014 Operator-carrying overhead cranes. Operator-carrying overhead cranes...

  10. 30 CFR 57.16014 - Operator-carrying overhead cranes.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 30 Mineral Resources 1 2011-07-01 2011-07-01 false Operator-carrying overhead cranes. 57.16014 Section 57.16014 Mineral Resources MINE SAFETY AND HEALTH ADMINISTRATION, DEPARTMENT OF LABOR METAL AND... Storage and Handling § 57.16014 Operator-carrying overhead cranes. Operator-carrying overhead cranes...

  11. 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...

  12. 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...

  13. 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...

  14. 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...

  15. 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...

  16. 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...

  17. 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...

  18. 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...

  19. 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...

  20. 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...

  1. 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

  2. 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.

  3. Enhanced Malignant Tumorigenesis in Cdk4-Transgenic Mice

    PubMed Central

    Miliani de Marval, Paula L.; Macias, Everardo; Conti, Claudio J.; Rodriguez-Puebla, Marcelo L.

    2010-01-01

    In a previous study, we reported that overexpression of 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. PMID:14647432

  4. 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 Central

    Tang, Bin; Seredenina, Tamara; Coppola, Giovanni; Kuhn, Alexandre; Geschwind, Daniel H.; Luthi-Carter, Ruth; Thomas, Elizabeth A.

    2011-01-01

    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 (R6/2Q300 transgenic mice), those carrying ~150 CAG repeats (R6/2Q150 transgenic mice) and littermate wt 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 R6/2Q300 transgenic mice. Upregulated genes in the R6/2Q300 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 R6/2Q300 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 R6/2Q300 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 R6/2Q300 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 R6/2Q150 transgenic mice. We suggest that the

  5. 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.

  6. Transgenic swine for biomedicine and agriculture.

    PubMed

    Prather, R S; Hawley, R J; Carter, D B; Lai, L; Greenstein, J L

    2003-01-01

    Initial technologies for creating transgenic swine only permitted random integration of the construct. However, by combining the technology for homologous recombination in fetal somatic cells with that of nuclear transfer (NT), it is now possible to create specific modifications to the swine genome. The first such example is that of knocking out a gene that is responsible for hyperacute rejection (HAR) when organs from swine are transferred to primates. Because swine are widely used as models of human diseases, there are opportunities for genetic modification to alter these models or to create additional models of human disease. Unfortunately, some of the offspring resulting from NT have abnormal phenotypes. However, it appears that these abnormal phenotypes are a result of epigenetic modifications and, thus, are not transmitted to the offspring of the clones. Although the technique of producing animals with specific genetic modifications by NT has been achieved, improvements to the NT technique as well as improvements in the culture conditions for somatic cells and the techniques for genetic modification are still needed.

  7. Detection of Transgenes on DNA Fibers.

    PubMed

    Shibata, Fukashi

    2016-01-01

    Fluorescence in situ hybridization (FISH) was developed for detecting specific DNA sequences directly on mitotic or meiotic chromosomes. However, the resolution of FISH on chromosomes is limited by condensed structure of chromatin, and it is difficult to differentiate two target sites close to each other. To overcome this issue, the objects was changed to stretched DNA fibers, and this fiber FISH technique has now been used for revealing genome structure at molecular level. Hybridization and detection procedures of fiber FISH are common with FISH on chromosomes. Therefore, application of fiber FISH is not difficult for the researchers of some experience in ordinary FISH. DNA fibers can be released from nuclei fixed on glass slides using a detergent. The DNA fibers were shred in FISH procedure, and the resultant fragments became small bead-like shape. This makes FISH signals on DNA fibers a series of dots. The size of DNA in the dot is estimated to be approximately 1 kb, it corresponding to the resolution of fiber FISH. This makes it possible to analyze structures of transgenes on DNA fibers in detail. PMID:27557695

  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. 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.

  10. Cardiac-Targeted Transgenic Mutant Mitochondrial Enzymes

    PubMed Central

    Kohler, James J.; Hosseini, Seyed H.; Green, Elgin; Hoying-Brandt, Amy; Cucoranu, Ioan; Haase, Chad P.; Russ, Rodney; Srivastava, Jaya; Ivey, Kristopher; Ludaway, Tomika; Kapoor, Victor; Abuin, Allison; Shapoval, Alexsey; Santoianni, Robert; Saada, Ann; Elpeleg, Orly; Lewis, William

    2009-01-01

    Mitochondrial (mt) DNA biogenesis is critical to cardiac contractility. DNA polymerase gamma (pol γ) replicates mtDNA, whereas thymidine kinase 2 (TK2) monophosphorylates pyrimidines intramitochondrially. Point mutations in POLG and TK2 result in clinical diseases associated with mtDNA depletion and organ dysfunction. Pyrimidine analogs (NRTIs) inhibit Pol γ and mtDNA replication. Cardiac “dominant negative” murine transgenes (TGs; Pol γ Y955G, and TK2 H121N or I212N) defined the role of each in the heart. mtDNA abundance, histopathological features, histochemistry, mitochondrial protein abundance, morphometry, and echocardiography were determined for TGs in “2 × 2” studies with or without pyrimidine analogs. Cardiac mtDNA abundance decreased in Y955C TGs (∼50%) but increased in H121N and I212N TGs (20-70%). Succinate dehydrogenase (SDH) increased in hearts of all mutants. Ultrastructural changes occurred in Y955C and H121N TGs. Histopathology demonstrated hypertrophy in H121N, LV dilation in I212N, and both hypertrophy and dilation in Y955C TGs. Antiretrovirals increased LV mass (≈50%) for all three TGs which combined with dilation indicates cardiomyopathy. Taken together, these studies demonstrate three manifestations of cardiac dysfunction that depend on the nature of the specific mutation and antiretroviral treatment. Mutations in genes for mtDNA biogenesis increase risk for defective mtDNA replication, leading to LV hypertrophy. PMID:18446447

  11. Production of diabetic offspring using cryopreserved epididymal sperm by in vitro fertilization and intrafallopian insemination techniques in transgenic pigs.

    PubMed

    Umeyama, Kazuhiro; Honda, Kasumi; Matsunari, Hitomi; Nakano, Kazuaki; Hidaka, Tatsuro; Sekiguchi, Keito; Mochizuki, Hironori; Takeuchi, Yasuhiro; Fujiwara, Tsukasa; Watanabe, Masahito; Nagaya, Masaki; Nagashima, Hiroshi

    2013-12-17

    Somatic cell nuclear transfer (SCNT) is a useful technique for creating pig strains that model human diseases. However, production of numerous cloned disease model pigs by SCNT for large-scale experiments is impractical due to its complexity and inefficiency. In the present study, we aimed to establish an efficient procedure for proliferating the diabetes model pig carrying the mutant human hepatocyte nuclear factor-1α gene. A founder diabetes transgenic cloned pig was generated by SCNT and treated with insulin to allow for normal growth to maturity, at which point epididymal sperm could be collected for cryopreservation. In vitro fertilization and intrafallopian insemination using the cryopreserved epididymal sperm resulted in diabetes model transgenic offspring. These results suggest that artificial reproductive technology using cryopreserved epididymal sperm could be a practical option for proliferation of genetically modified disease model pigs. PMID:23979397

  12. Production of Diabetic Offspring Using Cryopreserved Epididymal Sperm by In Vitro Fertilization and Intrafallopian Insemination Techniques in Transgenic Pigs

    PubMed Central

    UMEYAMA, Kazuhiro; HONDA, Kasumi; MATSUNARI, Hitomi; NAKANO, Kazuaki; HIDAKA, Tatsuro; SEKIGUCHI, Keito; MOCHIZUKI, Hironori; TAKEUCHI, Yasuhiro; FUJIWARA, Tsukasa; WATANABE, Masahito; NAGAYA, Masaki; NAGASHIMA, Hiroshi

    2013-01-01

    Abstract Somatic cell nuclear transfer (SCNT) is a useful technique for creating pig strains that model human diseases. However, production of numerous cloned disease model pigs by SCNT for large-scale experiments is impractical due to its complexity and inefficiency. In the present study, we aimed to establish an efficient procedure for proliferating the diabetes model pig carrying the mutant human hepatocyte nuclear factor-1α gene. A founder diabetes transgenic cloned pig was generated by SCNT and treated with insulin to allow for normal growth to maturity, at which point epididymal sperm could be collected for cryopreservation. In vitro fertilization and intrafallopian insemination using the cryopreserved epididymal sperm resulted in diabetes model transgenic offspring. These results suggest that artificial reproductive technology using cryopreserved epididymal sperm could be a practical option for proliferation of genetically modified disease model pigs. PMID:23979397

  13. Receptor editing in a transgenic mouse model: site, efficiency, and role in B cell tolerance and antibody diversification.

    PubMed

    Pelanda, R; Schwers, S; Sonoda, E; Torres, R M; Nemazee, D; Rajewsky, K

    1997-12-01

    Mice carrying transgenic rearranged V region genes in their IgH and Igkappa loci to encode an autoreactive specificity direct the emerging autoreactive progenitors into a pre-B cell compartment, in which their receptors are edited by secondary Vkappa-Jkappa rearrangements and RS recombination. Editing is an efficient process, because the mutant mice generate normal numbers of B cells. In a similar nonautoreactive transgenic strain, neither a pre-B cell compartment nor receptor editing was seen. Thus, the pre-B cell compartment may have evolved to edit the receptors of autoreactive cells and later been generally exploited for efficient antibody diversification through the invention of the pre-B cell receptor, mimicking an autoreactive antibody to direct the bulk of the progenitors into that compartment.

  14. 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.

  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. PMID:25371551

  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. RNAi-mediated knockdown of IKK1 in transgenic mice using a transgenic construct containing the human H1 promoter.

    PubMed

    Moreno-Maldonado, Rodolfo; Murillas, Rodolfo; Navarro, Manuel; Page, Angustias; Suarez-Cabrera, Cristian; Alameda, Josefa P; Bravo, Ana; Casanova, M Llanos; Ramirez, Angel

    2014-01-01

    Inhibition of gene expression through siRNAs is a tool increasingly used for the study of gene function in model systems, including transgenic mice. To achieve perdurable effects, the stable expression of siRNAs by an integrated transgenic construct is necessary. For transgenic siRNA expression, promoters transcribed by either RNApol II or III (such as U6 or H1 promoters) can be used. Relatively large amounts of small RNAs synthesis are achieved when using RNApol III promoters, which can be advantageous in knockdown experiments. To study the feasibility of H1 promoter-driven RNAi-expressing constructs for protein knockdown in transgenic mice, we chose IKK1 as the target gene. Our results indicate that constructs containing the H1 promoter are sensitive to the presence of prokaryotic sequences and to transgene position effects, similar to RNApol II promoters-driven constructs. We observed variable expression levels of transgenic siRNA among different tissues and animals and a reduction of up to 80% in IKK1 expression. Furthermore, IKK1 knockdown led to hair follicle alterations. In summary, we show that constructs directed by the H1 promoter can be used for knockdown of genes of interest in different organs and for the generation of animal models complementary to knockout and overexpression models. PMID:24523631

  18. 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.

  19. Diabetes-Associated Dry Eye Syndrome in a New Humanized Transgenic Model of Type 1 Diabetes

    PubMed Central

    Imam, Shahnawaz; Elagin, Raya B.

    2013-01-01

    Purpose Patients with Type 1 Diabetes (T1D) are at high risk of developing lacrimal gland dysfunction. We have developed a new model of human T1D using double-transgenic mice carrying HLA-DQ8 diabetes-susceptibility haplotype instead of mouse MHC-class II and expressing the human beta cell autoantigen Glutamic Acid Decarboxylase in pancreatic beta cells. We report here the development of dry eye syndrome (DES) after diabetes induction in our humanized transgenic model. Methods Double-transgenic mice were immunized with DNA encoding human GAD65, either naked or in adenoviral vectors, to induce T1D. Mice monitored for development of diabetes developed lacrimal gland dysfunction. Results Animals developed lacrimal gland disease (classically associated with diabetes in Non Obese Diabetic [NOD] mice and with T1D in humans) as they developed glucose intolerance and diabetes. Animals manifested obvious clinical signs of dry eye syndrome (DES), from corneal erosions to severe keratitis. Histological studies of peri-bulbar areas revealed lymphocytic infiltration of glandular structures. Indeed, infiltrative lesions were observed in lacrimal/Harderian glands within weeks following development of glucose intolerance. Lesions ranged from focal lymphocytic infiltration to complete acinar destruction. We observed a correlation between the severity of the pancreatic infiltration and the severity of the ocular disease. Conclusions Our results demonstrate development of DES in association with antigen-specific insulitis and diabetes following immunization with clinically relevant human autoantigen concomitantly expressed in pancreatic beta cells of diabetes-susceptible mice. As in the NOD mouse model and as in human T1D, our animals developed diabetes-associated DES. This specific finding stresses the relevance of our model for studying these human diseases. We believe our model will facilitate studies to prevent/treat diabetes-associated DES as well as human diabetes. PMID

  20. Accumulation of functional recombinant human coagulation factor IX in transgenic soybean seeds.

    PubMed

    Cunha, Nicolau B; Murad, André M; Ramos, Gustavo L; Maranhão, Andréia Q; Brígido, Marcelo M; Araújo, Ana Cláudia G; Lacorte, Cristiano; Aragão, Francisco J L; Covas, Dimas T; Fontes, Aparecida M; Souza, Gustavo H M F; Vianna, Giovanni R; Rech, Elíbio L

    2011-08-01

    The seed-based production of recombinant proteins is an efficient strategy to achieve the accumulation, correct folding, and increased stability of these recombinant proteins. Among potential plant molecular farming systems, soybean [Glycine max (L.) Merrill] is a viable option for the production of recombinant proteins due to its high protein content, known regulatory sequences, efficient gene transfer protocols, and a scalable production system under greenhouse conditions. We report here the expression and stable accumulation of human coagulation factor IX (hFIX) in transgenic soybean seeds. A biolistic process was utilised to co-introduce a plasmid carrying the hFIX gene under the transcriptional control of the α' subunit of a β-conglycinin seed-specific promoter and an α-Coixin signal peptide in soybean embryonic axes from mature seeds. The 56-kDa hFIX protein was expressed in the transgenic seeds at levels of up to 0.23% (0.8 g kg(-1) seed) of the total soluble seed protein as determined by an enzyme-linked immunosorbent assay (ELISA) and western blot. Ultrastructural immunocytochemistry assays indicated that the recombinant hFIX in seed cotyledonary cells was efficiently directed to protein storage vacuoles. Mass spectrometry characterisation confirmed the presence of the hFIX recombinant protein sequence. Protein extracts from transgenic seeds showed a blood-clotting activity of up to 1.4% of normal plasma. Our results demonstrate the correct processing and stable accumulation of functional hFIX in soybean seeds stored for 6 years under room temperature conditions (22 ± 2°C).

  1. Improved protein quality in transgenic soybean expressing a de novo synthetic protein, MB-16.

    PubMed

    Zhang, Yunfang; Schernthaner, Johann; Labbé, Natalie; Hefford, Mary A; Zhao, Jiping; Simmonds, Daina H

    2014-06-01

    To improve soybean [Glycine max (L.) Merrill] seed nutritional quality, a synthetic gene, MB-16 was introduced into the soybean genome to boost seed methionine content. MB-16, an 11 kDa de novo protein enriched in the essential amino acids (EAAs) methionine, threonine, lysine and leucine, was originally developed for expression in rumen bacteria. For efficient seed expression, constructs were designed using the soybean codon bias, with and without the KDEL ER retention sequence, and β-conglycinin or cruciferin seed specific protein storage promoters. Homozygous lines, with single locus integrations, were identified for several transgenic events. Transgene transmission and MB-16 protein expression were confirmed to the T5 and T7 generations, respectively. Quantitative RT-PCR analysis of developing seed showed that the transcript peaked in growing seed, 5-6 mm long, remained at this peak level to the full-sized green seed and then was significantly reduced in maturing yellow seed. Transformed events carrying constructs with the rumen bacteria codon preference showed the same transcription pattern as those with the soybean codon preference, but the transcript levels were lower at each developmental stage. MB-16 protein levels, as determined by immunoblots, were highest in full-sized green seed but the protein virtually disappeared in mature seed. However, amino acid analysis of mature seed, in the best transgenic line, showed a significant increase of 16.2 and 65.9 % in methionine and cysteine, respectively, as compared to the parent. This indicates that MB-16 elevated the sulfur amino acids, improved the EAA seed profile and confirms that a de novo synthetic gene can enhance the nutritional quality of soybean.

  2. Enhancing lignan biosynthesis by over-expressing pinoresinol lariciresinol reductase in transgenic wheat.

    PubMed

    Ayella, Allan K; Trick, Harold N; Wang, Weiqun

    2007-12-01

    Lignans are phenylpropane dimers that are biosynthesized via the phenylpropanoid pathway, in which pinoresinol lariciresinol reductase (PLR) catalyzes the last steps of lignan production. Our previous studies demonstrated that the contents of lignans in various wheat cultivars were significantly associated with anti-tumor activities in APC(Min) mice. To enhance lignan biosynthesis, this study was conducted to transform wheat cultivars ('Bobwhite', 'Madison', and 'Fielder', respectively) with the Forsythia intermedia PLR gene under the regulatory control of maize ubiquitin promoter. Of 24 putative transgenic wheat lines, we successfully obtained 3 transformants with the inserted ubiquitin-PLR gene as screened by PCR. Southern blot analysis further demonstrated that different copies of the PLR gene up to 5 were carried out in their genomes. Furthermore, a real-time PCR indicated approximately 17% increase of PLR gene expression over the control in 2 of the 3 positive transformants at T(0) generation. The levels of secoisolariciresinol diglucoside, a prominent lignan in wheat as determined by HPLC-MS, were found to be 2.2-times higher in one of the three positive transgenic sub-lines at T(2 )than that in the wild-type (117.9 +/- 4.5 vs. 52.9 +/- 19.8 mug/g, p <0.005). To the best of our knowledge, this is the first study that elevated lignan levels in a transgenic wheat line has been successfully achieved through genetic engineering of over-expressed PLR gene. Although future studies are needed for a stably expression and more efficient transformants, the new wheat line with significantly higher SDG contents obtained from this study may have potential application in providing additive health benefits for cancer prevention.

  3. 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

  4. [Determination of nutrient elements in transgenic insect-resistant cotton tissues by three different spectroscopical methods].

    PubMed

    Sun, Cai-Xia; Zhang, Yu-Lan; Sun, Yu-Quan; Yang, Lei; Wang, Jie; Cui, Zhen-Bo

    2009-11-01

    In order to find out the effects of exogenous genes, such as Bt and Bt coupled with CpTI, on nutrition metabolism in transgenic plants, totally eleven types of nutrient elements in transgenic Bt (Z30) and Bt-CpTI (CCRI41 and SGK321) cotton were determined using methods of flame atomic absorption spectroscopy, flame atomic emission spectroscopy and spectrophotometry at flowering stage and boll-opening stage. The results showed that the chemical composition of plant nutrition in transgenic insect-resistant cotton differed in comparison with non-transgenic cotton counterparts related to varieties, tissues and stages. The content of total N in transgenic cotton changed most significantly. Especially, it increased by 21% for transgenic Bt cotton Z30 compared to non-transgenic cotton Z16. These changes in total N content were probably caused by both transgenes expression in transgenic cotton and other processes not studied in this experiment. The content of Mg, Na and Cu in transgenic cotton varied significantly only in some certain varieties or tissues. It was unobvious how the incorporation of transgenes impacted on the content of organic C, total P, total S, K, Ca, Fe and Zn in transgenic cotton. The authors speculated that there were no significant changes in utilization and accumulation of these nutrient elements between transgenic insect-resistant cotton and their non-transgenic cotton counterparts (Z16, CCRI23 and SY321, respectively).

  5. Differential regulation of the expression in transgenic tobacco of the gene for beta-glucuronidase under the control of the 5'-upstream regions of two catalase genes from castor bean.

    PubMed

    Suzuki, M; Miyamoto, R; Hattori, T; Nakamura, K; Asahi, T

    1995-03-01

    The regulatory functions of the 5'-flanking regions of two genes for catalase (cat1 and cat2) from castor bean were analyzed in transgenic tobacco plants that carried fusion constructs that included the gene for beta-glucuronidase (GUS) for Escherichia coli. Dry mature seeds from transgenic plants carrying the CAT1-GUS or CAT2-GUS constructs, in which the GUS gene was fused to the 5'-flanking region of cat1 or cat2, respectively, contained significant GUS activity, indicating that the promoters of cat1 and cat2 were active during seed development. GUS activity increased in response to germination in the seeds of transgenic tobacco that carried CAT1-GUS, as well as in those that carried CAT2-GUS. During the post-germinative stage the GUS activity directed by CAT2-GUS increased still further, whereas that directed by CAT1-GUS decreased. The changes in GUS activity in the transgenic tobacco plants that carried CAT1-GUS and CAT2-GUS were similar to the changes in the levels of transcripts of cat1 and cat2, respectively, in castor bean. The results suggest that the expression of cat1 and cat2 in the germinating seeds and post-germinative seedlings is regulated mainly at the level of transcription. However, the distribution of GUS activity among the organs of the transgenic tobacco seedlings and plantlets, which was examined by histochemical staining and by enzymatic assays of tissue extracts, was not identical to that of transcripts of cat1 and cat2 in castor bean. Histochemical analysis also revealed the interesting spatial regulation of the expression of the promoter of cat2 in the transgenic tobacco seedlings.

  6. Additive transgene expression and genetic introgression in multiple green-fluorescent protein transgenic crop x weed hybrid generations.

    PubMed

    Halfhill, M D; Millwood, R J; Weissinger, A K; Warwick, S I; Stewart, C N

    2003-11-01

    The level of transgene expression in crop x weed hybrids and the degree to which crop-specific genes are integrated into hybrid populations are important factors in assessing the potential ecological and agricultural risks of gene flow associated with genetic engineering. The average transgene zygosity and genetic structure of transgenic hybrid populations change with the progression of generations, and the green fluorescent protein (GFP) transgene is an ideal marker to quantify transgene expression in advancing populations. The homozygous T(1) single-locus insert GFP/ Bacillus thuringiensis (Bt) transgenic canola ( Brassica napus, cv Westar) with two copies of the transgene fluoresced twice as much as hemizygous individuals with only one copy of the transgene. These data indicate that the expression of the GFP gene was additive, and fluorescence could be used to determine zygosity status. Several hybrid generations (BC(1)F(1), BC(2)F(1)) were produced by backcrossing various GFP/Bt transgenic canola ( B. napus, cv Westar) and birdseed rape ( Brassica rapa) hybrid generations onto B. rapa. Intercrossed generations (BC(2)F(2) Bulk) were generated by crossing BC(2)F(1) individuals in the presence of a pollinating insect ( Musca domestica L.). The ploidy of plants in the BC(2)F(2) Bulk hybrid generation was identical to the weedy parental species, B. rapa. AFLP analysis was used to quantify the degree of B. napus introgression into multiple backcross hybrid generations with B. rapa. The F(1) hybrid generations contained 95-97% of the B. napus-specific AFLP markers, and each successive backcross generation demonstrated a reduction of markers resulting in the 15-29% presence in the BC(2)F(2) Bulk population. Average fluorescence of each successive hybrid generation was analyzed, and homozygous canola lines and hybrid populations that contained individuals homozygous for GFP (BC(2)F(2) Bulk) demonstrated significantly higher fluorescence than hemizygous hybrid

  7. Additive transgene expression and genetic introgression in multiple green-fluorescent protein transgenic crop x weed hybrid generations.

    PubMed

    Halfhill, M D; Millwood, R J; Weissinger, A K; Warwick, S I; Stewart, C N

    2003-11-01

    The level of transgene expression in crop x weed hybrids and the degree to which crop-specific genes are integrated into hybrid populations are important factors in assessing the potential ecological and agricultural risks of gene flow associated with genetic engineering. The average transgene zygosity and genetic structure of transgenic hybrid populations change with the progression of generations, and the green fluorescent protein (GFP) transgene is an ideal marker to quantify transgene expression in advancing populations. The homozygous T(1) single-locus insert GFP/ Bacillus thuringiensis (Bt) transgenic canola ( Brassica napus, cv Westar) with two copies of the transgene fluoresced twice as much as hemizygous individuals with only one copy of the transgene. These data indicate that the expression of the GFP gene was additive, and fluorescence could be used to determine zygosity status. Several hybrid generations (BC(1)F(1), BC(2)F(1)) were produced by backcrossing various GFP/Bt transgenic canola ( B. napus, cv Westar) and birdseed rape ( Brassica rapa) hybrid generations onto B. rapa. Intercrossed generations (BC(2)F(2) Bulk) were generated by crossing BC(2)F(1) individuals in the presence of a pollinating insect ( Musca domestica L.). The ploidy of plants in the BC(2)F(2) Bulk hybrid generation was identical to the weedy parental species, B. rapa. AFLP analysis was used to quantify the degree of B. napus introgression into multiple backcross hybrid generations with B. rapa. The F(1) hybrid generations contained 95-97% of the B. napus-specific AFLP markers, and each successive backcross generation demonstrated a reduction of markers resulting in the 15-29% presence in the BC(2)F(2) Bulk population. Average fluorescence of each successive hybrid generation was analyzed, and homozygous canola lines and hybrid populations that contained individuals homozygous for GFP (BC(2)F(2) Bulk) demonstrated significantly higher fluorescence than hemizygous hybrid

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

    PubMed

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

    2010-12-01

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

  9. 25 CFR 23.51 - Grant carry-over authority.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 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...

  10. 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...

  11. 25 CFR 23.51 - Grant carry-over authority.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 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...

  12. 25 CFR 23.51 - Grant carry-over authority.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 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. High expression of human beta S- and alpha-globins in transgenic mice: erythrocyte abnormalities, organ damage, and the effect of hypoxia.

    PubMed Central

    Fabry, M E; Costantini, F; Pachnis, A; Suzuka, S M; Bank, N; Aynedjian, H S; Factor, S M; Nagel, R L

    1992-01-01

    A line of transgenic mice with two cointegrated transgenes, the human beta S- and alpha 2-globin genes, linked to the beta-globin locus control region was produced and bred with mice carrying a deletion of the mouse beta major-globin gene. In transgenic mice homozygous for the beta major deletion (alpha H beta S[beta MDD]; where alpha H is human alpha-globin and MD is mouse deletion), 72.5 +/- 2.4% (mean +/- SD) of the beta-chains are beta S and the ratio of alpha H- to beta S-globin was 0.73. Introduction of a heterozygous mouse alpha-globin deletion into mice homozygous for the beta major deletion (alpha H beta S[alpha MD beta MDD]) resulted in 65.1 +/- 8.5% beta S and a human alpha/beta ratio of 0.89 +/- 0.2. Sickling occurs in 95% of erythrocytes from alpha H beta S[beta MDD] mice after slow deoxygenation. Transmission electron microscopy revealed polymer fiber formation but not fascicles of fiber. Increased organ weight was noted in lung, spleen, and kidney of transgenic mice vs. controls that may be due to hypertrophy or increased blood volume in the lungs and/or increased tissue water content. The hemoglobin content of lung, spleen, and kidney was also elevated in transgenic animals due to trapped hemoglobin and/or increased blood volume. When transgenic and control mice were examined by magnetic resonance imaging at 9.4 tesla, some transgenic animals had enlarged kidneys with prolonged relaxation time, consistent with increased organ weight and water content. The glomerular filtration rate was elevated in transgenic animals, which is characteristic of young sickle cell patients. Furthermore, exposure to hypoxia resulted in significantly decreased hematocrit, increased erythrocyte density, and induced a urine-concentrating defect. We conclude that the transgenic mouse line reported here has chronic organ damage and further hematological and organ dysfunction can be induced by hypoxia. Images PMID:1465455

  14. Detailed characterization of Mirafiori lettuce virus-resistant transgenic lettuce.

    PubMed

    Kawazu, Yoichi; Fujiyama, Ryoi; Noguchi, Yuji; Kubota, Masaharu; Ito, Hidekazu; Fukuoka, Hiroyuki

    2010-04-01

    Lettuce big-vein disease is caused by Mirafiori lettuce virus (MiLV), which is vectored by the soil-borne fungus Olpidium brassicae. A MiLV-resistant transgenic lettuce line was developed through introducing inverted repeats of the MiLV coat protein (CP) gene. Here, a detailed characterization study of this lettuce line was conducted by comparing it with the parental, non-transformed 'Kaiser' cultivar. There were no significant differences between transgenic and non-transgenic lettuce in terms of pollen fertility, pollen dispersal, seed production, seed dispersal, dormancy, germination, growth of seedlings under low or high temperature, chromatographic patterns of leaf extracts, or effects of lettuce on the growth of broccoli or soil microflora. A significant difference in pollen size was noted, but the difference was small. The length of the cotyledons of the transgenic lettuce was shorter than that of 'Kaiser,' but there were no differences in other morphological characteristics. Agrobacterium tumefaciens used for the production of transgenic lettuce was not detected in transgenic seeds. The transgenic T(3), T(4), and T(5) generations showed higher resistance to MiLV and big-vein symptoms expression than the resistant 'Pacific' cultivar, indicating that high resistance to lettuce big-vein disease is stably inherited. PCR analysis showed that segregation of the CP gene was nearly 3:1 in the T(1) and T(2) generations, and that the transgenic T(3) generation was homozygous for the CP gene. Segregation of the neomycin phosphotransferase II (npt II) gene was about 3:1 in the T(1) generation, but the full length npt II gene was not detected in the T(2) or T(3) generation. The segregation pattern of the CP and npt II genes in the T(1) generation showed the expected 9:3:3:1 ratio. These results suggest that the fragment including the CP gene and that including the npt II gene have been integrated into two unlinked loci, and that the T(1) plant selected in our study did

  15. Development of transgenic watermelon resistant to Cucumber mosaic virus and Watermelon mosaic virus by using a single chimeric transgene construct.

    PubMed

    Lin, Ching-Yi; Ku, Hsin-Mei; Chiang, Yi-Hua; Ho, Hsiu-Yin; Yu, Tsong-Ann; Jan, Fuh-Jyh

    2012-10-01

    Watermelon, an important fruit crop worldwide, is prone to attack by several viruses that often results in destructive yield loss. To develop a transgenic watermelon resistant to multiple virus infection, a single chimeric transgene comprising a silencer DNA from the partial N gene of Watermelon silver mottle virus (WSMoV) fused to the partial coat protein (CP) gene sequences of Cucumber mosaic virus (CMV), Cucumber green mottle mosaic virus (CGMMV) and Watermelon mosaic virus (WMV) was constructed and transformed into watermelon (cv. Feeling) via Agrobacterium-mediated transformation. Single or multiple transgene copies randomly inserted into various locations in the genome were confirmed by Southern blot analysis. Transgenic watermelon R(0) plants were individually challenged with CMV, CGMMV or WMV, or with a mixture of these three viruses for resistance evaluation. Two lines were identified to exhibit resistance to CMV, CGMMV, WMV individually, and a mixed inoculation of the three viruses. The R(1) progeny of the two resistant R(0) lines showed resistance to CMV and WMV, but not to CGMMV. Low level accumulation of transgene transcripts in resistant plants and small interfering (si) RNAs specific to CMV and WMV were readily detected in the resistant R(1) plants by northern blot analysis, indicating that the resistance was established via RNA-mediated post-transcriptional gene silencing (PTGS). Loss of the CGMMV CP-transgene fragment in R1 progeny might be the reason for the failure to resistant CGMMV infection, as shown by the absence of a hybridization signal and no detectable siRNA specific to CGMMV in Southern and northern blot analyses. In summary, this study demonstrated that fusion of different viral CP gene fragments in transgenic watermelon contributed to multiple virus resistance via PTGS. The construct and resistant watermelon lines developed in this study could be used in a watermelon breeding program for resistance to multiple viruses.

  16. Immunity to tomato yellow leaf curl virus in transgenic tomato is associated with accumulation of transgene small RNA.

    PubMed

    Leibman, Diana; Prakash, Shanmugam; Wolf, Dalia; Zelcer, Aaron; Anfoka, Ghandi; Haviv, Sabrina; Brumin, Marina; Gaba, Victor; Arazi, Tzahi; Lapidot, Moshe; Gal-On, Amit

    2015-11-01

    Gene silencing is a natural defense response of plants against invading RNA and DNA viruses. The RNA post-transcriptional silencing system has been commonly utilized to generate transgenic crop plants that are "immune" to plant virus infection. Here, we applied this approach against the devastating DNA virus tomato yellow leaf curl virus (TYLCV) in its host tomato (Solanum lycopersicum L.). To generate broad resistance to a number of different TYLCV viruses, three conserved sequences (the intergenic region [NCR], V1-V2 and C1-C2 genes) from the genome of the severe virus (TYLCV) were synthesized as a single insert and cloned into a hairpin configuration in a binary vector, which was used to transform TYLCV-susceptible tomato plants. Eight of 28 independent transgenic tomato lines exhibited immunity to TYLCV-Is and to TYLCV-Mld, but not to tomato yellow leaf curl Sardinia virus, which shares relatively low sequence homology with the transgene. In addition, a marker-free (nptII-deleted) transgenic tomato line was generated for the first time by Agrobacterium-mediated transformation without antibiotic selection, followed by screening of 1180 regenerated shoots by whitefly-mediated TYLCV inoculation. Resistant lines showed a high level of transgene-siRNA (t-siRNA) accumulation (22% of total small RNA) with dominant sizes of 21 nt (73%) and 22 nt (22%). The t-siRNA displayed hot-spot distribution ("peaks") along the transgene, with different distribution patterns than the viral-siRNA peaks observed in TYLCV-infected tomato. A grafting experiment demonstrated the mobility of 0.04% of the t-siRNA from transgenic rootstock to non-transformed scion, even though scion resistance against TYLCV was not achieved. PMID:26255053

  17. The Dmp1-SOST Transgene Interacts With and Downregulates the Dmp1-Cre Transgene and the Rosa(Notch) Allele.

    PubMed

    Zanotti, Stefano; Canalis, Ernesto

    2016-05-01

    Activation of Notch1 in osteocytes of Rosa(Notch) mice, where a loxP-flanked STOP cassette and the Nicd coding sequence were targeted to the reverse orientation splice acceptor (Rosa)26 locus, causes osteopetrosis associated with suppressed Sost expression and enhanced Wnt signaling. To determine whether Sost downregulation mediates the effects of Notch activation in osteocytes, Rosa(Notch) mice were crossed with transgenics expressing Cre recombinase or SOST under the control of the dentin matrix protein (Dmp)1 promoter. Dmp1-SOST transgenics displayed vertebral osteopenia and a modest femoral cancellous and cortical bone phenotype, whereas hemizygous Dmp1-Cre transgenics heterozygous for the Rosa(Notch) allele (Dmp1-Cre;Rosa(Notch)) exhibited osteopetrosis. The phenotype of Notch activation in osteocytes was prevented in Dmp1-Cre;Rosa(Notch) mice hemizygous for the Dmp1-SOST transgene. The effect was associated with downregulated Notch signaling and suppressed Dmp1 and Rosa26 expression. To test whether SOST regulates Notch expression in osteocytes, cortical bone cultures from Dmp1-Cre;Rosa(Notch) mice or from Rosa(Notch) control littermates were exposed to recombinant human SOST. The addition of SOST had only modest effects on Notch target gene mRNA levels and suppressed Dmp1, but not Cre or Rosa26, expression. These findings suggest that prevention of the Dmp1-Cre;Rosa(Notch) skeletal phenotype by Dmp1-SOST is not secondary to SOST expression but to interactions among the Dmp1-SOST and Dmp1-Cre transgenes and the Rosa26 locus. In conclusion, the Dmp1-SOST transgene suppresses the expression of the Dmp1-Cre transgene and of Rosa26. PMID:26456319

  18. 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.

  19. 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

  20. Antibiotic-resistant soil bacteria in transgenic plant fields

    PubMed Central

    Demanèche, Sandrine; Sanguin, Hervé; Poté, John; Navarro, Elisabeth; Bernillon, Dominique; Mavingui, Patrick; Wildi, Walter; Vogel, Timothy M.; Simonet, Pascal

    2008-01-01

    Understanding the prevalence and polymorphism of antibiotic resistance genes in soil bacteria and their potential to be transferred horizontally is required to evaluate the likelihood and ecological (and possibly clinical) consequences of the transfer of these genes from transgenic plants to soil bacteria. In this study, we combined culture-dependent and -independent approaches to study the prevalence and diversity of bla genes in soil bacteria and the potential impact that a 10-successive-year culture of the transgenic Bt176 corn, which has a blaTEM marker gene, could have had on the soil bacterial community. The bla gene encoding resistance to ampicillin belongs to the beta-lactam antibiotic family, which is widely used in medicine but is readily compromised by bacterial antibiotic resistance. Our results indicate that soil bacteria are naturally resistant to a broad spectrum of beta-lactam antibiotics, including the third cephalosporin generation, which has a slightly stronger discriminating effect on soil isolates than other cephalosporins. These high resistance levels for a wide range of antibiotics are partly due to the polymorphism of bla genes, which occur frequently among soil bacteria. The blaTEM116 gene of the transgenic corn Bt176 investigated here is among those frequently found, thus reducing any risk of introducing a new bacterial resistance trait from the transgenic material. In addition, no significant differences were observed in bacterial antibiotic-resistance levels between transgenic and nontransgenic corn fields, although the bacterial populations were different. PMID:18292221

  1. 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

  2. Targeting gene expression to the wool follicle in transgenic sheep.

    PubMed

    Damak, S; Jay, N P; Barrell, G K; Bullock, D W

    1996-02-01

    To establish the feasibility of overexpressing foreign genes in the wool follicle, transgenic sheep were produced by pronuclear microinjection of a DNA construct consisting of a mouse ultrahigh-sulfur keratin promoter linked to the bacterial chloramphenicol acetyl transferase (CAT) gene. Four of 31 lambs born were transgenic. The overall efficiency of transgenesis was 1.1% of zygotes injected and transferred. Two transgenic rams were mated to nontransgenic ewes, and both transmitted the gene to their offspring in Mendelian fashion. CAT expression was found in the skin of one G0 ram and in 9 out of 26 transgenic G1 progeny. Two G1 lambs were sacrificed to study tissue specificity. Both had high levels of expression in skin but One had high expression in spleen and kidney with lower levels of expression in lung; the other had low expression in spleen, lung, and muscle. In situ hybridization demonstrated that transgene expression in the skin was confined to the keratogenous zone of the wool follicle cortex. Expression of CAT activity in skin was correlated with diet-induced or seasonal changes in the rate of wool growth. This keratin promoter appears useful for overexpressing factors in the wool follicle that might influence wool production or properties.

  3. 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

  4. 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

    2016-01-01

    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. PMID:26864449

  5. Biodegradation of explosives by transgenic plants expressing pentaerythritol tetranitrate reductase.

    PubMed

    French, C E; Rosser, S J; Davies, G J; Nicklin, S; Bruce, N C

    1999-05-01

    Plants offer many advantages over bacteria as agents for bioremediation; however, they typically lack the degradative capabilities of specially selected bacterial strains. Transgenic plants expressing microbial degradative enzymes could combine the advantages of both systems. To investigate this possibility in the context of bioremediation of explosive residues, we generated transgenic tobacco plants expressing pentaerythritol tetranitrate reductase, an enzyme derived from an explosive-degrading bacterium that enables degradation of nitrate ester and nitroaromatic explosives. Seeds from transgenic plants were able to germinate and grow in the presence of 1 mM glycerol trinitrate (GTN) or 0.05 mM trinitrotoluene, at concentrations that inhibited germination and growth of wild-type seeds. Transgenic seedlings grown in liquid medium with 1 mM GTN showed more rapid and complete denitration of GTN than wild-type seedlings. This example suggests that transgenic plants expressing microbial degradative genes may provide a generally applicable strategy for bioremediation of organic pollutants in soil. PMID:10331811

  6. Retinoic acid-mediated gene expression in transgenic reporter zebrafish.

    PubMed

    Perz-Edwards, A; Hardison, N L; Linney, E

    2001-01-01

    Retinoic acid-mediated gene activation is important for normal vertebrate development. The size and nature of retinoic acid make it difficult to identify the precise cellular location of this signaling molecule throughout an embryo. Additionally, retinoic acid (RA) signaling is regulated by a complex combination of receptors, coactivators, and antagonizing proteins. Thus, in order to integrate these signals and identify regions within a whole developing embryo where cells can respond transcriptionally to retinoic acid, we have used a reporter transgenic approach. We have generated several stable lines of transgenic zebrafish which use retinoic acid response elements to drive fluorescent protein expression. In these zebrafish lines, transgene expression is localized to regions of the neural tube, retina, notochord, somites, heart, pronephric ducts, branchial arches, and jaw muscles in embryos and larvae. Transgene expression can be induced in additional regions of the neural tube and retina as well as the immature notochord, hatching gland, enveloping cell layer, and fin by exposing embryos to retinoic acid. Treatment with retinoic acid synthase inhibitors, citral and diethylaminobenzaldehyde (DEAB), during neurulation, greatly reduces transgene expression. DEAB treatment of embryos at gastrulation phenocopies the embryonic effects of vitamin A deprivation or targeted disruption of the RA synthase retinaldehyde dehydrogenase-2 in other vertebrates. Together these data suggest that the reporter expression we see in zebrafish is dependent upon conserved vertebrate pathways of RA synthesis.

  7. [Improving the nutritional value of plant foods through transgenic approaches].

    PubMed

    Wu, Yong-Mei; Mao, Xue; Wang, Shu-Jian; Li, Run-Zhi

    2004-07-01

    The most nutrients required in the human diet come from plants. The nutritional quality of plant products affects the human healthy. The advance of molecular cloning and transgenic technology has provided a new way to enhance the nutritional value of plant material. Transgenic modification of plant nutritional value has progressed greatly in the following aspects: improving the quality, composition and levels of protein, starch and fatty acid in different crops; increasing the levels of antioxidants (e.g. carotenoids and flavonoids); breeding the new type of plants with medical value for human. To date, many transgenic plants with nutritional enhancement have been developed. These transgenic plant products could be directly used as human diet or as valued materials in developing the "functional food" with especial nutritional quality and healthy effects after they are approved by a series of evaluations on their safety and nutritional efficiency for human being. We designed new zinc finger transcription factors (ZFP-TFs) that can specifically down-regulate the expression of the endogenous soybean FAD2-1 gene which catalyzes oleic acid to linoleic acid. Seed-specific expression of these ZFP-TFs in transgenic soybean somatic embryos repressed FAD2-1 transcription and increased significantly the levels of oleic acid, indicating that the engineered ZFP-TFs are capable of regulating fatty acid metabolism and modulating the expression of endogenous genes in plants.

  8. 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.

  9. Loss of sense transgene-induced post-transcriptional gene silencing by sequential introduction of the same transgene sequences in tobacco.

    PubMed

    Hirai, Sayaka; Takahashi, Kouta; Abiko, Tomomi; Kodama, Hiroaki

    2010-04-01

    RNA silencing is an epigenetic inhibition of gene expression and is guided by small interfering RNAs. Sense transgene-induced post-transcriptional gene silencing (S-PTGS) occurs in a portion of a transgenic plant population. When a sense transgene encoding a tobacco endoplasmic reticulum omega-3 fatty acid desaturase (NtFAD3) was introduced into tobacco plants, an S-PTGS line, S44, was obtained. Introduction of another copy of the NtFAD3 transgene into S44 plants caused a phenotypic change from S-PTGS to overexpression. Because this change was associated with the methylation of the promoter sequences of the transgene, reduced transcriptional activity may abolish S-PTGS and residual transcription of the sense transgene may account for the overexpression. To clarify whether RNA-directed DNA methylation (RdDM) can repress the transcriptional activity of the S44 transgene locus, we introduced several RdDM constructs targeting the transgene promoter. An RdDM construct harboring a 200-bp-long fragment of promoter sequences efficiently abrogated the generation of NtFAD3 small interfering RNAs in S44 plants. Transcription of the transgene was partially repressed, but the resulting NtFAD3 mRNAs successfully accumulated and an overexpressed phenotype was established. Our results indicate an example in which overexpression of the transgene is established by complex epigenetic interactions among the transgenic loci. PMID:20180844

  10. 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.

  11. 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

    2012-01-01

    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. PMID:22869542

  12. Pollen-mediated gene flow from transgenic cotton under greenhouse conditions is dependent on different pollinators.

    PubMed

    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

  13. Dissection of the locus control function located on the chicken lysozyme gene domain in transgenic mice.

    PubMed Central

    Bonifer, C; Yannoutsos, N; Krüger, G; Grosveld, F; Sippel, A E

    1994-01-01

    The entire chicken lysozyme gene locus including all known cis-regulatory sequences and the 5' and 3' matrix attachment sites defining the borders of the DNase I sensitive chromatin domain, is expressed at a high level and independent of its chromosomal position in macrophages of transgenic mice. It was concluded that the lysozyme gene locus carries a locus control function. We analysed several cis-regulatory deletion mutants to investigate their influence on tissue specificity and level of expression. Position independent expression of the gene is lost whenever one of the upstream tissue specific enhancer regions is deleted, although tissue specific expression is usually retained. Deletion of the domain border fragments has no influence on copy number dependency of expression. However, without these regions an increased incidence of ectopic expression is observed. This suggests that the domain border fragments may help to suppress transgene expression in inappropriate tissues. We conclude, that position independent expression of the lysozyme gene is not controlled by a single specific region of the locus but is the result of the concerted action of several tissue specific upstream regulatory DNA elements with the promoter. Images PMID:7937146

  14. Impaired glucose homeostasis in transgenic mice expressing the human transient neonatal diabetes mellitus locus, TNDM

    PubMed Central

    Ma, Dan; Shield, Julian P.H.; Dean, Wendy; Leclerc, Isabelle; Knauf, Claude; Burcelin, Rémy; Rutter, Guy A.; Kelsey, Gavin

    2004-01-01

    Transient neonatal diabetes mellitus (TNDM) is a rare inherited diabetic syndrome apparent in the first weeks of life and again during early adulthood. The relative contributions of reduced islet β cell number and impaired β cell function to the observed hypoinsulinemia are unclear. The inheritance pattern of this imprinted disorder implicates overexpression of one or both genes within the TNDM locus: ZAC, which encodes a proapoptotic zinc finger protein, and HYMAI, which encodes an untranslated mRNA. To investigate the consequences for pancreatic function, we have developed a high-copy transgenic mouse line, TNDM29, carrying the human TNDM locus. TNDM29 neonates display hyperglycemia, and older adults, impaired glucose tolerance. Neonatal hyperglycemia occurs only on paternal transmission, analogous to paternal dependence of TNDM in humans. Embryonic pancreata of TNDM29 mice showed reductions in expression of endocrine differentiation factors and numbers of insulin-staining structures. By contrast, β cell mass was normal or elevated at all postnatal stages, whereas pancreatic insulin content in neonates and peak serum insulin levels after glucose infusion in adults were reduced. Expression of human ZAC and HYMAI in these transgenic mice thus recapitulates key features of TNDM and implicates impaired development of the endocrine pancreas and β cell function in disease pathogenesis. PMID:15286800

  15. Stress tolerance of transgenic barley accumulating the alfalfa aldose reductase in the cytoplasm and the chloroplast.

    PubMed

    Nagy, Bettina; Majer, Petra; Mihály, Róbert; Pauk, János; Horváth, Gábor V

    2016-09-01

    Barley represents one of the major crops grown worldwide; its genetic transformation provides an important tool for the improvement of crop quality and tolerance to environmental stress factors. Biotic and abiotic stresses produce reactive oxygen species in the plant cells that can directly oxidize the cellular components including lipid membranes; resulting in lipid peroxidation and subsequently the accumulation of reactive carbonyl compounds. In order to protect barley plants from the effects of stress-produced reactive carbonyls, an Agrobacterium-mediated transformation was carried out using the Medicago sativa aldose reductase (MsALR) gene. In certain transgenic lines the produced MsALR enzyme was targeted to the chloroplasts to evaluate its protective effect in these organelles. The dual fluorescent protein-based method was used for the evaluation of tolerance of young seedlings to diverse stresses; our results demonstrated that this technique could be reliably applied for the detection of cellular stress in a variety of conditions. The chlorophyll and carotenoid content measurements also supported the results of the fluorescent protein-based method and the stress-protective effect of the MsALR enzyme. Targeting of MsALR into the chloroplast has also resulted in increased stress tolerance, similarly to the observed effect of the cytosolic MsALR accumulation. The results of the DsRed/GFP fluorescent protein-based method indicated that both the cytosol and chloroplast accumulation of MsALR can increase the abiotic stress tolerance of transgenic barley lines. PMID:27469099

  16. 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. PMID:27162494

  17. Pollen-mediated gene flow from transgenic cotton under greenhouse conditions is dependent on different pollinators.

    PubMed

    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.

  18. Evaluation of transgenic Bt corn for resistance to the Asian corn borer (Lepidoptera: Pyralidae).

    PubMed

    He, Kanglai; Wang, Zhenying; Zhou, Darong; Wen, Liping; Song, Yanying; Yao, Zhuyun

    2003-06-01

    The Asian corn borer, Ostrinia furnacalis (Guenée), is the most important insect pest on corn in China. Bt transgenic corn provides a new tool for Asian corn borer control. Monsanto's YieldGard Bt transgenic corn expressing Cry1Ab protein, and a non-Bt control, were evaluated in Beijing. Laboratory bioassays were carried out by exposing neonates to an agar-free diet containing Bt corn whorl leaves, tassels, and anthers, or by exposing neonates directly to fresh silk and pollen. These are the tissues initially attacked by neonates in the field. All of these tissues, with the exception of pollen, contained sufficient insecticidal protein to kill > or = 95% of larvae within 7 d. Surviving larvae had also not grown beyond first instar and weighed < or = 0.1 mg. Although larvae feeding on Bt corn pollen were significantly smaller than those on non-Bt corn pollen, there was no significant difference in mortality. Field trials were also conducted with artificial infestations of Asian corn borer at mid whorl, late whorl, and silking stages. Damage ratings and number of larvae surviving per plant indicated that Bt corn was highly resistant to Asian corn borer. Therefore, YieldGard offers the potential for season-long protection against first- and second-generation Asian corn borer.

  19. 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

  20. Modulation of inflammation in transgenic models of Alzheimer’s disease

    PubMed Central

    2014-01-01

    Over the past decade the process of inflammation has been a focus of increasing interest in the Alzheimer’s disease (AD) field, not only for its potential role in neuronal degeneration but also as a promising therapeutic target. However, recent research in this field has provided divergent outcomes, largely due to the use of different models and different stages of the disease when the investigations have been carried out. It is now accepted that microglia, and possibly astrocytes, change their activation phenotype during ageing and the stage of the disease, and therefore these are important factors to have in mind to define the function of different inflammatory components as well as potential therapies. Modulating inflammation using animal models of AD has offered the possibility to investigate inflammatory components individually and manipulate inflammatory genes in amyloid precursor protein and tau transgenics independently. This has also offered some hints on the mechanisms by which these factors may affect AD pathology. In this review we examine the different transgenic approaches and treatments that have been reported to modulate inflammation using animal models of AD. These studies have provided evidence that enhancing inflammation is linked with increases in amyloid-beta (Aβ) generation, Aβ aggregation and tau phosphorylation. However, the alterations on tau phosphorylation can be independent of changes in Aβ levels by these inflammatory mediators. PMID:24490742

  1. Generation of transgene-free induced pluripotent mouse stem cells by the piggyBac transposon

    PubMed Central

    Yusa, Kosuke; Rad, Roland; Takeda, Junji; Bradley, Allan

    2009-01-01

    Induced pluripotent stem cells (iPSCs) have been generated from somatic cells by transgenic expression of Oct4, Sox2, Klf4, and cMyc. A major difficulty in the application of this technology for regenerative medicine, however, is the delivery of reprogramming factors. Whereas retroviral transduction increases the risk of tumorigenicity, transient expression methods have considerably lower reprogramming efficiencies. Here we show a highly efficient piggyBac transposon-based approach to generate integration-free iPSCs. Transposons carrying 2A peptide-linked reprogramming factors induced reprogramming of mouse embryonic fibroblasts with equivalent efficiencies to retroviral transduction. Transposons were removed from these primary iPSCs by re-expressing transposase. Transgene-free iPSCs could be easily identified by HSVtk-FIAU selection. piggyBac excises without a footprint, leaving the iPSC genome without any genetic alteration. iPSCs fulfilled all criteria of pluripotency, such as expression of embryonic stem cell-specific markers, teratoma formation and contribution to chimeras. piggyBac transposon-based reprogramming may be used to generate therapeutically applicable iPSCs. PMID:19337237

  2. 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.

  3. The transposition frequency of Tag1 elements is increased in transgenic Arabidopsis lines.

    PubMed Central

    Bhatt, A M; Lister, C; Crawford, N; Dean, C

    1998-01-01

    Tag1 was identified as a highly active endogenous transposable element in transgenic Arabidopsis thaliana Landsberg erecta plants carrying the maize transposable element Activator (Ac). Here, we describe experiments designed to determine the basis for the high activity of Tag1. The frequency of transposition of Tag1 elements was compared in lines containing or lacking Ac transposase to assess the effect of Ac transposase on Tag1 activity. Three populations of nontransgenic plants, including nontransformed regenerants, were also analyzed. The high level of activity of Tag1 did not correlate with the presence or absence of Ac transposase but was significantly higher in transgenic lines. This result was maintained through at least six generations after transformation. These data suggest that Tag1 transposition is stimulated by processes that occur during the Agrobacterium transformation and that thereafter remain active. Two Tag1 elements are tightly linked in the Landsberg erecta genome and map to the lower arm of chromosome 1. Tag1 elements were found in only a few A. thaliana ecotypes but were present in four other Arabidopsis species. PMID:9501115

  4. BrUGE1 transgenic rice showed improved growth performance with enhanced drought tolerance

    PubMed Central

    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

    2016-01-01

    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. PMID:27162494

  5. 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.

  6. Evaluation of transgenic Bt corn for resistance to the Asian corn borer (Lepidoptera: Pyralidae).

    PubMed

    He, Kanglai; Wang, Zhenying; Zhou, Darong; Wen, Liping; Song, Yanying; Yao, Zhuyun

    2003-06-01

    The Asian corn borer, Ostrinia furnacalis (Guenée), is the most important insect pest on corn in China. Bt transgenic corn provides a new tool for Asian corn borer control. Monsanto's YieldGard Bt transgenic corn expressing Cry1Ab protein, and a non-Bt control, were evaluated in Beijing. Laboratory bioassays were carried out by exposing neonates to an agar-free diet containing Bt corn whorl leaves, tassels, and anthers, or by exposing neonates directly to fresh silk and pollen. These are the tissues initially attacked by neonates in the field. All of these tissues, with the exception of pollen, contained sufficient insecticidal protein to kill > or = 95% of larvae within 7 d. Surviving larvae had also not grown beyond first instar and weighed < or = 0.1 mg. Although larvae feeding on Bt corn pollen were significantly smaller than those on non-Bt corn pollen, there was no significant difference in mortality. Field trials were also conducted with artificial infestations of Asian corn borer at mid whorl, late whorl, and silking stages. Damage ratings and number of larvae surviving per plant indicated that Bt corn was highly resistant to Asian corn borer. Therefore, YieldGard offers the potential for season-long protection against first- and second-generation Asian corn borer. PMID:12852639

  7. 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... hazardous materials on passenger-carrying vehicles, exceptions. No hazardous materials except small-arms ammunition, emergency shipments of drugs, chemicals and hospital supplies, and the accompanying munitions...

  8. 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... hazardous materials on passenger-carrying vehicles, exceptions. No hazardous materials except small-arms ammunition, emergency shipments of drugs, chemicals and hospital supplies, and the accompanying munitions...

  9. 49 CFR 177.870 - Regulations for passenger carrying vehicles.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... PUBLIC HIGHWAY Regulations Applying to Hazardous Material on Motor Vehicles Carrying Passengers for Hire... hazardous materials on passenger-carrying vehicles, exceptions. No hazardous materials except small-arms ammunition, emergency shipments of drugs, chemicals and hospital supplies, and the accompanying munitions...

  10. 49 CFR 177.870 - Regulations for passenger carrying vehicles.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... PUBLIC HIGHWAY Regulations Applying to Hazardous Material on Motor Vehicles Carrying Passengers for Hire... hazardous materials on passenger-carrying vehicles, exceptions. No hazardous materials except small-arms ammunition, emergency shipments of drugs, chemicals and hospital supplies, and the accompanying munitions...

  11. 49 CFR 177.870 - Regulations for passenger carrying vehicles.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... PUBLIC HIGHWAY Regulations Applying to Hazardous Material on Motor Vehicles Carrying Passengers for Hire... hazardous materials on passenger-carrying vehicles, exceptions. No hazardous materials except small-arms ammunition, emergency shipments of drugs, chemicals and hospital supplies, and the accompanying munitions...

  12. 14 CFR 91.869 - Carry-forward compliance.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 14 Aeronautics and Space 2 2014-01-01 2014-01-01 false Carry-forward compliance. 91.869 Section 91...) AIR TRAFFIC AND GENERAL OPERATING RULES GENERAL OPERATING AND FLIGHT RULES Operating Noise Limits § 91.869 Carry-forward compliance. (a) Any operator that exceeds the requirements of paragraph (b) of §...

  13. 14 CFR 91.869 - Carry-forward compliance.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 14 Aeronautics and Space 2 2013-01-01 2013-01-01 false Carry-forward compliance. 91.869 Section 91...) AIR TRAFFIC AND GENERAL OPERATING RULES GENERAL OPERATING AND FLIGHT RULES Operating Noise Limits § 91.869 Carry-forward compliance. (a) Any operator that exceeds the requirements of paragraph (b) of §...

  14. 14 CFR 91.869 - Carry-forward compliance.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 14 Aeronautics and Space 2 2012-01-01 2012-01-01 false Carry-forward compliance. 91.869 Section 91...) AIR TRAFFIC AND GENERAL OPERATING RULES GENERAL OPERATING AND FLIGHT RULES Operating Noise Limits § 91.869 Carry-forward compliance. (a) Any operator that exceeds the requirements of paragraph (b) of §...

  15. 14 CFR 91.869 - Carry-forward compliance.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 14 Aeronautics and Space 2 2010-01-01 2010-01-01 false Carry-forward compliance. 91.869 Section 91...) AIR TRAFFIC AND GENERAL OPERATING RULES GENERAL OPERATING AND FLIGHT RULES Operating Noise Limits § 91.869 Carry-forward compliance. (a) Any operator that exceeds the requirements of paragraph (b) of §...

  16. 14 CFR 91.869 - Carry-forward compliance.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 14 Aeronautics and Space 2 2011-01-01 2011-01-01 false Carry-forward compliance. 91.869 Section 91...) AIR TRAFFIC AND GENERAL OPERATING RULES GENERAL OPERATING AND FLIGHT RULES Operating Noise Limits § 91.869 Carry-forward compliance. (a) Any operator that exceeds the requirements of paragraph (b) of §...

  17. 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...

  18. 46 CFR 122.340 - Vessels carrying vehicles.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 46 Shipping 4 2011-10-01 2011-10-01 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 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...

  20. 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…