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Sample records for gene transfer systems

  1. Novel gene transfer systems: intelligent gene transfer vectors for gene medicines.

    PubMed

    Nakajima, Toshihiro

    2012-01-01

    Drug delivery systems for gene transfer are called 'vectors'. These systems were originally invented as a delivery system for the transfection in vitro or in vivo. Several vectors are then developed for clinical use of gene medicines and currently some of them are approved as animal drugs. Conventional drug delivery system generally consists of approved (existing) materials to avoid additional pre-clinical or clinical studies. However, current vectors contain novel materials to improve an efficacy of gene medicines. Thus, these vectors have functions more than a mere delivery of active ingredients. For example some vectors have immunological functions such as adjuvants in vaccines. These new types of vectors are called 'intelligent' or 'innovative' vector system', since the concept or strategy for the development is completely different from conventional drug delivery systems. In this article, we described a current status of 'intelligent gene transfer vectors and discussed on the potentials of them.

  2. Methods for Gene Transfer to the Central Nervous System

    PubMed Central

    Kantor, Boris; Bailey, Rachel M.; Wimberly, Keon; Kalburgi, Sahana N.; Gray, Steven J.

    2015-01-01

    Gene transfer is an increasingly utilized approach for research and clinical applications involving the central nervous system (CNS). Vectors for gene transfer can be as simple as an unmodified plasmid, but more commonly involve complex modifications to viruses to make them suitable gene delivery vehicles. This chapter will explain how tools for CNS gene transfer have been derived from naturally occurring viruses. The current capabilities of plasmid, retroviral, adeno-associated virus, adenovirus, and herpes simplex virus vectors for CNS gene delivery will be described. These include both focal and global CNS gene transfer strategies, with short- or long-term gene expression. As is described in this chapter, an important aspect of any vector is the cis-acting regulatory elements incorporated into the vector genome that control when, where, and how the transgene is expressed. PMID:25311922

  3. Effects of local and systemic viral interleukin-10 gene transfer on corneal allograft survival.

    PubMed

    Gong, N; Pleyer, U; Volk, H-D; Ritter, T

    2007-03-01

    In this study, we explored the immunomodulatory effects of viral interleukin (IL) IL-10 after ex vivo and in vivo gene transfer in experimental corneal transplantation. Wistar-Furth rats were used as donors and major histocompatibility complex class I/II-disparate Lewis rats served as recipients. For ex vivo gene therapy donor corneas were either transfected with liposome/vIL-10 plasmid DNA mixtures or transduced with a vIL-10 expressing adenovirus vector (AdvIL-10). For in vivo studies, recipients were treated with AdvIL-10 intraperitoneally 1 day before transplantation. Graft survival was analysed using the Kaplan-Meier survival method. To monitor the efficacy of the therapy messenger RNA (mRNA) cytokine expression profiles in grafts and draining lymph nodes were analysed by quantitative real-time reverse transcription-polymerase chain reaction. Moreover, anti-adenovirus immunity was also investigated. Neither ex vivo liposome-mediated vIL-10 gene transfer nor ex vivo AdvIL-10 gene transfer led to prolonged corneal allograft survival. In contrast, corneal allograft survival was significantly prolonged in animals receiving systemic AdvIL-10 gene transfer. Moreover, only systemic vIL-10 gene therapy modulated the cytokine mRNA expression profile in draining lymph nodes. Interestingly, systemic AdvIL-10 gene transfer could not inhibit the generation of anti-adenovirus antibodies. Our data indicate systemic expression of the vIL-10 gene is required to modulate the cytokine expression profile in the draining lymph nodes, which might be a pre-requisite for the success of cytokine gene therapy.

  4. Systemic protein delivery by muscle-gene transfer is limited by a local immune response

    PubMed Central

    Wang, Lixin; Dobrzynski, Eric; Schlachterman, Alexander; Cao, Ou; Herzog, Roland W.

    2005-01-01

    Adeno-associated viral (AAV) vectors have been successfully used for therapeutic expression of systemic transgene products (such as factor IX or erythropoietin) following in vivo administration to skeletal muscle of animal models of inherited hematologic disorders. However, an immune response may be initiated if the transgene product represents a neoantigen. Here, we use ovalbumin (OVA) as a model antigen and demonstrate immune-mediated elimination of expression on muscle-directed AAV-2 gene transfer. Administration to immune competent mice resulted in transient systemic OVA expression. Within 10 days, OVA-specific T-helper cells had been activated in draining lymph nodes, an inflammatory immune response ensued, and OVA-expressing muscle fibers were destroyed by a cytotoxic CD8+ T-cell response. Use of a muscle-specific promoter did not prevent this immune response. Adoptively transferred CD4+ cells transgenic for a T-cell receptor specific to OVA peptide-major histocompatibility complex class II showed antigen-specific, vector dose-dependent proliferation confined to the draining lymph nodes of AAV-OVA–transduced muscle within 5 days after gene transfer and subsequently participated in lymphocytic infiltration of transduced muscle. This study documents that a local immune response limits sustained expression of a secreted protein in muscle gene transfer, a finding that may have consequences for design of clinical protocols. PMID:15713796

  5. Type IV secretion systems: tools of bacterial horizontal gene transfer and virulence

    PubMed Central

    Juhas, Mario; Crook, Derrick W; Hood, Derek W

    2008-01-01

    Type IV secretion systems (T4SSs) are multisubunit cell-envelope-spanning structures, ancestrally related to bacterial conjugation machines, which transfer proteins and nucleoprotein complexes across membranes. T4SSs mediate horizontal gene transfer, thus contributing to genome plasticity and the evolution of pathogens through dissemination of antibiotic resistance and virulence genes. Moreover, T4SSs are also used for the delivery of bacterial effector proteins across the bacterial membrane and the plasmatic membrane of eukaryotic host cell, thus contributing directly to pathogenicity. T4SSs are usually encoded by multiple genes organized into a single functional unit. Based on a number of features, the organization of genetic determinants, shared homologies and evolutionary relationships, T4SSs have been divided into several groups. Type F and P (type IVA) T4SSs resembling the archetypal VirB/VirD4 system of Agrobacterium tumefaciens are considered to be the paradigm of type IV secretion, while type I (type IVB) T4SSs are found in intracellular bacterial pathogens, Legionella pneumophila and Coxiella burnetii. Several novel T4SSs have been identified recently and their functions await investigation. The most recently described GI type T4SSs play a key role in the horizontal transfer of a wide variety of genomic islands derived from a broad spectrum of bacterial strains. PMID:18549454

  6. [Gene transfer system mediated by PEI-cholesterol lipopolymer with lipid microbubbles].

    PubMed

    Jiang, Yong-Nan; Mo, Hong-Ying; Chen, Jian-Hai

    2010-05-01

    The properties of polyethyleneimine-cholesterol cationic lipopolymer (PEI-Chol) as gene carries and its gene transfer efficiency in vitro with lipid microbubbles were presented in this paper. PEI-Chol lipopolymer was synthesized by linking cholesterol chloroformate to the amino groups of branched poly(ethyleneimine) (PEI) of 1 800. The structure and molecular weight of PEI-Chol were confirmed by IR, 1H NMR and MADI-TOF-MS (matrix-assisted laser desorption/ionization time-of-flight tandem mass spectrometry), respectively. The average molecular weight of PEI-Chol was approximately 2 000. The gene delivery system of bubble/PEI-Chol/DNA was constructed by mixed PEI-Chol/pDNA (N/P 10:1) complexes with lipid microbubbles (2-8 microm) which were prepared by DPPC, DSPE-PEG2000 and perfluoropropane with the reverse phase evaporation technique. pEGFP-Cl (enhanced green fluorescent protein) was used as report gene to investigate the DNA condensing ability of PEI-Chol lipopolymer by agarose gel electrophoresis. And their cytotoxicity and in vitro transfer efficiency of different complexes were compared with each other in A549 and MCF-7. The results indicated PEI-Chol lipopolymer can condense plasmid DNA when N/P ratio upto 4, PEI-Chol complexes and bubble/PEI-Chol/DNA complexes were nontoxic to A549 and MCF-7 when formulated at the N/P ratio of 10/1 as determined by MTT assay. This bubble/PEI-Chol/DNA delivery system provided good transfer efficiency with other desirable characteristics such as against-precipitation of plasma proteins. In conclusion, bubble/PEI-Chol/DNA complex is a novel non-viral gene delivery system.

  7. Widespread Central Nervous System Gene Transfer and Silencing After Systemic Delivery of Novel AAV-AS Vector.

    PubMed

    Choudhury, Sourav R; Harris, Anne F; Cabral, Damien J; Keeler, Allison M; Sapp, Ellen; Ferreira, Jennifer S; Gray-Edwards, Heather L; Johnson, Jacob A; Johnson, Aime K; Su, Qin; Stoica, Lorelei; DiFiglia, Marian; Aronin, Neil; Martin, Douglas R; Gao, Guangping; Sena-Esteves, Miguel

    2016-04-01

    Effective gene delivery to the central nervous system (CNS) is vital for development of novel gene therapies for neurological diseases. Adeno-associated virus (AAV) vectors have emerged as an effective platform for in vivo gene transfer, but overall neuronal transduction efficiency of vectors derived from naturally occurring AAV capsids after systemic administration is relatively low. Here, we investigated the possibility of improving CNS transduction of existing AAV capsids by genetically fusing peptides to the N-terminus of VP2 capsid protein. A novel vector AAV-AS, generated by the insertion of a poly-alanine peptide, is capable of extensive gene transfer throughout the CNS after systemic administration in adult mice. AAV-AS is 6- and 15-fold more efficient than AAV9 in spinal cord and cerebrum, respectively. The neuronal transduction profile varies across brain regions but is particularly high in the striatum where AAV-AS transduces 36% of striatal neurons. Widespread neuronal gene transfer was also documented in cat brain and spinal cord. A single intravenous injection of an AAV-AS vector encoding an artificial microRNA targeting huntingtin (Htt) resulted in 33-50% knockdown of Htt across multiple CNS structures in adult mice. This novel AAV-AS vector is a promising platform to develop new gene therapies for neurodegenerative disorders.

  8. Phylogenetic evidence for extensive horizontal gene transfer of type III secretion system genes among enterobacterial plant pathogens.

    PubMed

    Naum, Marianna; Brown, Eric W; Mason-Gamer, Roberta J

    2009-10-01

    This study uses sequences from four genes, which are involved in the formation of the type III secretion apparatus, to determine the role of horizontal gene transfer in the evolution of virulence genes for the enterobacterial plant pathogens. Sequences of Erwinia, Brenneria, Pectobacterium, Dickeya and Pantoea were compared (a) with one another, (b) with sequences of enterobacterial animal pathogens, and (c) with sequences of plant pathogenic gamma and beta proteobacteria, to evaluate probable paths of lateral exchange leading to the current distribution of virulence determinants among these micro-organisms. Phylogenies were reconstructed based on hrcC, hrcR, hrcJ and hrcV gene sequences using parsimony and maximum-likelihood algorithms. Virulence gene phylogenies were also compared with several housekeeping gene loci in order to evaluate patterns of lateral versus vertical acquisition. The resulting phylogenies suggest that multiple horizontal gene transfer events have occurred both within and among the enterobacterial plant pathogens and plant pathogenic gamma and beta proteobacteria. hrcJ sequences are the most similar, exhibiting anywhere from 2 to 50 % variation at the nucleotide level, with the highest degree of variation present between plant and animal pathogen sequences. hrcV sequences are conserved among plant and animal pathogens at the N terminus. The C-terminal domain is conserved only among the enterobacterial plant pathogens, as are the hrcC and hrcR sequences. Additionally, hrcJ and hrcV sequence phylogenies suggest that at least some type III secretion system virulence genes from enterobacterial plant pathogens are related more closely to those of the genus Pseudomonas, a conclusion neither supported nor refuted by hrcC or hrcR.

  9. Gene therapy of the central nervous system: general considerations on viral vectors for gene transfer into the brain.

    PubMed

    Serguera, C; Bemelmans, A-P

    2014-12-01

    The last decade has nourished strong doubts on the beneficial prospects of gene therapy for curing fatal diseases. However, this climate of reservation is currently being transcended by the publication of several successful clinical protocols, restoring confidence in the appropriateness of therapeutic gene transfer. A strong sign of this present enthusiasm for gene therapy by clinicians and industrials is the market approval of the therapeutic viral vector Glybera, the first commercial product in Europe of this class of drug. This new field of medicine is particularly attractive when considering therapies for a number of neurological disorders, most of which are desperately waiting for a satisfactory treatment. The central nervous system is indeed a very compliant organ where gene transfer can be stable and successful if provided through an appropriate strategy. The purpose of this review is to present the characteristics of the most efficient virus-derived vectors used by researchers and clinicians to genetically modify particular cell types or whole regions of the brain. In addition, we discuss major issues regarding side effects, such as genotoxicity and immune response associated to the use of these vectors.

  10. Recent Origin of the Methacrylate Redox System in Geobacter sulfurreducens AM-1 through Horizontal Gene Transfer.

    PubMed

    Arkhipova, Oksana V; Meer, Margarita V; Mikoulinskaia, Galina V; Zakharova, Marina V; Galushko, Alexander S; Akimenko, Vasilii K; Kondrashov, Fyodor A

    2015-01-01

    The origin and evolution of novel biochemical functions remains one of the key questions in molecular evolution. We study recently emerged methacrylate reductase function that is thought to have emerged in the last century and reported in Geobacter sulfurreducens strain AM-1. We report the sequence and study the evolution of the operon coding for the flavin-containing methacrylate reductase (Mrd) and tetraheme cytochrome с (Mcc) in the genome of G. sulfurreducens AM-1. Different types of signal peptides in functionally interlinked proteins Mrd and Mcc suggest a possible complex mechanism of biogenesis for chromoproteids of the methacrylate redox system. The homologs of the Mrd and Mcc sequence found in δ-Proteobacteria and Deferribacteres are also organized into an operon and their phylogenetic distribution suggested that these two genes tend to be horizontally transferred together. Specifically, the mrd and mcc genes from G. sulfurreducens AM-1 are not monophyletic with any of the homologs found in other Geobacter genomes. The acquisition of methacrylate reductase function by G. sulfurreducens AM-1 appears linked to a horizontal gene transfer event. However, the new function of the products of mrd and mcc may have evolved either prior or subsequent to their acquisition by G. sulfurreducens AM-1.

  11. Novel and effective gene transfer technique for study of vascular renin angiotensin system.

    PubMed Central

    Morishita, R; Gibbons, G H; Kaneda, Y; Ogihara, T; Dzau, V J

    1993-01-01

    Vascular renin angiotensin system (RAS) has been reported to exist in vascular wall. However, there is no direct evidence whether the vascular RAS per se can modulate growth of vascular smooth muscle cells (VSMC), because there is no suitable method to investigate the effect of endogenously produced vasoactive substances on growth of these cells. In this study, we transferred angiotensin-converting enzyme (ACE) and/or renin cDNAs into cultured VSMC using the efficient Sendai virus (hemagglutinating virus of Japan) liposome-mediated gene transfer method, to examine their relative roles in VSMC growth in vitro. Within 35 min or 6 h, the transfection of ACE cDNA into VSMC by hemagglutinating virus of Japan method resulted in a twofold higher ACE activity than control vector, whereas a cationic liposome (Lipofectin)-mediated method failed to show any effect. This in vitro system provided us with the opportunity to investigate the influence of endogenous vascular RAS on VSMC growth. Transfection of ACE or renin cDNA resulted in increased DNA and RNA synthesis, which was inhibited with the specific angiotensin II receptor antagonist (DuP 753: 10(-6) M). Angiotensin I added to ACE-transfected VSMC increased RNA synthesis in a dose-dependent manner. Cotransfection of renin and ACE cDNAs stimulated further RNA synthesis as compared to ACE or renin cDNA alone. These results showed that transfected components of RAS can modulate VSMC growth through the endogenous production of vascular angiotensin II, and that ACE as well as renin are rate limiting in determining the VSMC RAS activity. We conclude that the hemagglutinating virus of Japan liposome-mediated gene transfer technique provides a new and useful tool for study of endogenous vascular modulators such as vascular RAS. Images PMID:8390484

  12. Inferring Horizontal Gene Transfer

    PubMed Central

    Lassalle, Florent; Dessimoz, Christophe

    2015-01-01

    Horizontal or Lateral Gene Transfer (HGT or LGT) is the transmission of portions of genomic DNA between organisms through a process decoupled from vertical inheritance. In the presence of HGT events, different fragments of the genome are the result of different evolutionary histories. This can therefore complicate the investigations of evolutionary relatedness of lineages and species. Also, as HGT can bring into genomes radically different genotypes from distant lineages, or even new genes bearing new functions, it is a major source of phenotypic innovation and a mechanism of niche adaptation. For example, of particular relevance to human health is the lateral transfer of antibiotic resistance and pathogenicity determinants, leading to the emergence of pathogenic lineages [1]. Computational identification of HGT events relies upon the investigation of sequence composition or evolutionary history of genes. Sequence composition-based ("parametric") methods search for deviations from the genomic average, whereas evolutionary history-based ("phylogenetic") approaches identify genes whose evolutionary history significantly differs from that of the host species. The evaluation and benchmarking of HGT inference methods typically rely upon simulated genomes, for which the true history is known. On real data, different methods tend to infer different HGT events, and as a result it can be difficult to ascertain all but simple and clear-cut HGT events. PMID:26020646

  13. Gene transfer in hepatocarcinoma cell lines: in vitro optimization of a virus-free system.

    PubMed

    Ghoumari, A M; Rixe, O; Yarovoi, S V; Zerrouqi, A; Mouawad, R; Poynard, T; Opolon, P; Khayat, D; Soubrane, C

    1996-06-01

    Many approaches exist for hepatic gene delivery, including viral vectors and non-viral vectors. In this study, we tested a panel of liposomes to transfer pAGO, a plasmid containing one copy of herpes simplex virus (HSVtk) gene, and pYED11, a plasmid containing two copies of the HSVtk gene, into a murine hepatocarcinoma cell line (Hepa 1-6) and a human hepatocarcinoma cell line (Hep-G2). The efficiency of gene delivery and expression was characterized by beta-galactosidase staining, flow cytometric analysis and quantitative lacZ activity. Different combinations of liposomes and DNA and the ratio of the concentration of liposome to DNA were tested. The efficient transfer was shown with DOTAP followed by transfectam and lipofectamine. Under these conditions, we tested the cytotoxicity of ganciclovir (GCV) exposure on Hepa 1-6 and Hep-G2 transfected separately with liposome-pAGO and liposome-pYED11 complexes. This study demonstrates the in vitro efficacy of each liposome tested to transduce the HSVtk gene into hepatocarcinoma cell lines. The transfer of two copies of the HSVtk gene rendered cells 1.5 times more sensitive to GCV than cells transduced by pAGO as compared to controls. This was achieved most efficiently by the DOTAP-pYED11 complex. Thus, pYED11 may be considered as an alternative to pAGO as a gene transfer vector.

  14. Molecular evidence for ongoing complementarity and horizontal gene transfer in endosymbiotic systems of mealybugs

    PubMed Central

    López-Madrigal, Sergio; Beltrà, Aleixandre; Resurrección, Serena; Soto, Antonia; Latorre, Amparo; Moya, Andrés; Gil, Rosario

    2014-01-01

    Intracellular bacterial supply of essential amino acids is common among sap-feeding insects, thus complementing the scarcity of nitrogenous compounds in plant phloem. This is also the role of the two mealybug endosymbiotic systems whose genomes have been sequenced. In the nested endosymbiotic system from Planococcus citri (Pseudococcinae), “Candidatus Tremblaya princeps” and “Candidatus Moranella endobia” cooperate to synthesize essential amino acids, while in Phenacoccus avenae (Phenacoccinae) this function is performed by its single endosymbiont “Candidatus Tremblaya phenacola.” However, little is known regarding the evolution of essential amino acid supplementation strategies in other mealybug systems. To address this knowledge gap, we screened for the presence of six selected loci involved in essential amino acid biosynthesis in five additional mealybug species. We found evidence of ongoing complementarity among endosymbionts from insects of subfamily Pseudococcinae, as well as horizontal gene transfer affecting endosymbionts from insects of family Phenacoccinae, providing a more comprehensive picture of the evolutionary history of these endosymbiotic systems. Additionally, we report two diagnostic motifs to help identify invasive mealybug species. PMID:25206351

  15. Systemic hyperosmolality improves beta-glucuronidase distribution and pathology in murine MPS VII brain following intraventricular gene transfer.

    PubMed

    Ghodsi, A; Stein, C; Derksen, T; Martins, I; Anderson, R D; Davidson, B L

    1999-11-01

    Mucopolysaccharidosis VII, a classical lysosomal storage disease, is caused by deficiency of the enzyme beta-glucuronidase. Central nervous system (CNS) manifestations are severe with accumulations of storage vacuoles in all cell types. Intraventricular gene transfer can lead to transduction of the ependyma, with production and secretion of beta-glucuronidase into the cerebral spinal fluid and underlying cortex resulting in reversal of disease pathology restricted to the periventricular areas. We tested if systemic hyperosmolality would increase the distribution of beta-glucuronidase in brain parenchyma after intraventricular virus injection. Mice were administered mannitol, intraperitoneally, 20 days after gene transfer and 1 day prior to sacrifice. Mannitol-induced systemic hyperosmolality caused a marked penetration of beta-glucuronidase into the brain parenchyma. If mannitol was administered at the time of the intraventricular injection of virus, there was penetration of vector across the ependymal cell layer, with infection of cells in the subependymal region. This also resulted in increased beta-glucuronidase activity throughout the brain. Sections of brains from beta-glucuronidase-deficient mice showed correction of cellular pathology in the subependymal region plus cortical structures away from the ventricular wall. These data indicate that virus-mediated gene transfer to the brain via the ventricles, coupled with systemic mannitol administration, can lead to extensive CNS distribution of beta-glucuronidase with concomitant correction of the storage defect. Our findings have positive therapeutic implications for the treatment of CNS disorders with gene transfer vectors and recombinant proteins.

  16. Lateral gene transfer, rearrangement, reconciliation

    PubMed Central

    2013-01-01

    Background Models of ancestral gene order reconstruction have progressively integrated different evolutionary patterns and processes such as unequal gene content, gene duplications, and implicitly sequence evolution via reconciled gene trees. These models have so far ignored lateral gene transfer, even though in unicellular organisms it can have an important confounding effect, and can be a rich source of information on the function of genes through the detection of transfers of clusters of genes. Result We report an algorithm together with its implementation, DeCoLT, that reconstructs ancestral genome organization based on reconciled gene trees which summarize information on sequence evolution, gene origination, duplication, loss, and lateral transfer. DeCoLT optimizes in polynomial time on the number of rearrangements, computed as the number of gains and breakages of adjacencies between pairs of genes. We apply DeCoLT to 1099 gene families from 36 cyanobacteria genomes. Conclusion DeCoLT is able to reconstruct adjacencies in 35 ancestral bacterial genomes with a thousand gene families in a few hours, and detects clusters of co-transferred genes. DeCoLT may also be used with any relationship between genes instead of adjacencies, to reconstruct ancestral interactions, functions or complexes. Availability http://pbil.univ-lyon1.fr/software/DeCoLT/ PMID:24564205

  17. Recent tissue engineering-based advances for effective rAAV-mediated gene transfer in the musculoskeletal system.

    PubMed

    Rey-Rico, Ana; Cucchiarini, Magali

    2016-04-01

    Musculoskeletal tissues are diverse and significantly different in their ability to repair upon injury. Current treatments often fail to reproduce the natural functions of the native tissue, leading to an imperfect healing. Gene therapy might improve the repair of tissues by providing a temporarily and spatially defined expression of the therapeutic gene(s) at the site of the injury. Several gene transfer vehicles have been developed to modify various human cells and tissues from musculoskeletal system among which the non-pathogenic, effective, and relatively safe recombinant adeno-associated viral (rAAV) vectors that have emerged as the preferred gene delivery system to treat human disorders. Adapting tissue engineering platforms to gene transfer approaches mediated by rAAV vectors is an attractive tool to circumvent both the limitations of the current therapeutic options to promote an effective healing of the tissue and the natural obstacles from these clinically adapted vectors to achieve an efficient and durable gene expression of the therapeutic sequences within the lesions.

  18. Three-layered polyplex micelle as a multifunctional nanocarrier platform for light-induced systemic gene transfer

    NASA Astrophysics Data System (ADS)

    Nomoto, Takahiro; Fukushima, Shigeto; Kumagai, Michiaki; Machitani, Kaori; Arnida; Matsumoto, Yu; Oba, Makoto; Miyata, Kanjiro; Osada, Kensuke; Nishiyama, Nobuhiro; Kataoka, Kazunori

    2014-04-01

    Nanocarriers responding to light have great potential for pinpoint therapy, and recent studies have revealed promising in vivo activity. However, light-selective gene transfer still remains challenging in the systemic application. Here we report systemic light-responsive nanocarriers for gene delivery developed through the sequential self-assembly of ABC-type triblock copolymer/DNA/dendrimeric photosensitizer, forming polyplex micelles with three-layered functional nanocompartments. The DNA-packaged core is covered by the photosensitizer-incorporated intermediate layer, which is encompassed by an outer shielding shell. This three-layered structure permits multistep photosensitizer and DNA delivery into a solid tumour by a systemic route: the shielding layer minimizes unfavourable interactions with blood components, and the photosensitizer is delivered to endo-/lysosomal membranes to facilitate light-selective cytoplasmic translocation of the micelles, accomplishing DNA delivery into the nucleus to exert gene expression. The polyplex micelles display >100-fold photoenhanced gene expression in cultured cells and exhibit light-induced in vivo gene transfer in solid tumours following systemic administration.

  19. Transfer system

    DOEpatents

    Kurosawa, Kanji; Koga, Bunichiro; Ito, Hideki; Kiriyama, Shigeru; Higuchi, Shizuo

    2003-05-20

    A transport system includes a traveling rail (1) which constitutes a transport route and a transport body (3) which is capable of traveling on the traveling rail in the longitudinal direction of the traveling rail. Flexible drive tubes (5) are arranged on the traveling rail in the longitudinal direction of the traveling rail. The transport body includes a traveling wheel (4) which is capable of rolling on the traveling rail and drive wheels (2) which are capable of rolling on the drive tubes upon receiving the rotational drive power generated by pressure of a pressure medium supplied to the drive tubes while depressing the drive tubes. The traveling rail includes a plurality of transport sections and the transport body is capable of receiving a rotational drive force from the drive tubes at every transport sections. If necessary, a transport route changeover switch which changes over the transport route can be provided between the transport sections.

  20. Systemic gene transfer reveals distinctive muscle transduction profile of tyrosine mutant AAV-1, -6, and -9 in neonatal dogs.

    PubMed

    Hakim, Chady H; Yue, Yongping; Shin, Jin-Hong; Williams, Regina R; Zhang, Keqing; Smith, Bruce F; Duan, Dongsheng

    2014-03-05

    The muscular dystrophies are a group of devastating genetic disorders that affect both skeletal and cardiac muscle. An effective gene therapy for these diseases requires bodywide muscle delivery. Tyrosine mutant adeno-associated virus (AAV) has been considered as a class of highly potent gene transfer vectors. Here, we tested the hypothesis that systemic delivery of tyrosine mutant AAV can result in bodywide muscle transduction in newborn dogs. Three tyrosine mutant AAV vectors (Y445F/Y731F AAV-1, Y445F AAV-6, and Y731F AAV-9) were evaluated. These vectors expressed the alkaline phosphatase reporter gene under transcriptional regulation of either the muscle-specific Spc5-12 promoter or the ubiquitous Rous sarcoma virus promoter. Robust skeletal and cardiac muscle transduction was achieved with Y445F/Y731F AAV-1. However, Y731F AAV-9 only transduced skeletal muscle. Surprisingly, Y445F AAV-6 resulted in minimal muscle transduction. Serological study suggests that the preexisting neutralization antibody may underlie the limited transduction of Y445F AAV-6. In summary, we have identified Y445F/Y731F AAV-1 as a potentially excellent systemic gene transfer vehicle to target both skeletal muscle and the heart in neonatal puppies. Our findings have important implications in exploring systemic neonatal gene therapy in canine models of muscular dystrophy.

  1. A Complete Set of Flagellar Genes Acquired by Horizontal Transfer Coexists with the Endogenous Flagellar System in Rhodobacter sphaeroides▿ †

    PubMed Central

    Poggio, Sebastian; Abreu-Goodger, Cei; Fabela, Salvador; Osorio, Aurora; Dreyfus, Georges; Vinuesa, Pablo; Camarena, Laura

    2007-01-01

    Bacteria swim in liquid environments by means of a complex rotating structure known as the flagellum. Approximately 40 proteins are required for the assembly and functionality of this structure. Rhodobacter sphaeroides has two flagellar systems. One of these systems has been shown to be functional and is required for the synthesis of the well-characterized single subpolar flagellum, while the other was found only after the genome sequence of this bacterium was completed. In this work we found that the second flagellar system of R. sphaeroides can be expressed and produces a functional flagellum. In many bacteria with two flagellar systems, one is required for swimming, while the other allows movement in denser environments by producing a large number of flagella over the entire cell surface. In contrast, the second flagellar system of R. sphaeroides produces polar flagella that are required for swimming. Expression of the second set of flagellar genes seems to be positively regulated under anaerobic growth conditions. Phylogenic analysis suggests that the flagellar system that was initially characterized was in fact acquired by horizontal transfer from a γ-proteobacterium, while the second flagellar system contains the native genes. Interestingly, other α-proteobacteria closely related to R. sphaeroides have also acquired a set of flagellar genes similar to the set found in R. sphaeroides, suggesting that a common ancestor received this gene cluster. PMID:17293429

  2. Panspermia and horizontal gene transfer

    NASA Astrophysics Data System (ADS)

    Klyce, Brig

    2009-08-01

    Evidence that extremophiles are hardy and ubiquitous is helping to make panspermia a respectable theory. But even if life on Earth originally came from space, biologists assume that the subsequent evolution of life is still governed by the darwinian paradigm. In this review we show how panspermia could amend darwinism and point to a cosmic source for, not only extremophiles but, all of life. This version of panspermia can be called "strong panspermia." To support this theory we will discuss recent evidence pertaining to horizontal gene transfer, viruses, genes apparently older than the Earthly evolution of the features they encode, and primate-specific genes without identifiable precursors.

  3. Inhibition of experimental autoimmune uveoretinitis by systemic and subconjunctival adenovirus-mediated transfer of the viral IL-10 gene

    PubMed Central

    De Kozak, Y; Thillaye-Goldenberg, B; Naud, M -C; Viana Da Costa, A; Auriault, C; Verwaerde, C

    2002-01-01

    Pathological ocular manifestations result from a dysregulation in the balance between proinflammatory type 1 cytokines and regulatory type 2 cytokines. Interleukin-10 (IL-10) is an anti-inflammatory cytokine with potent immunosuppressive effects. We have examined the efficiency of viral IL-10 adenovirus (Ad-vIL-10)-mediated gene transfer on experimental autoimmune uveoretinitis (EAU) induced in mice and rats by purified retinal autoantigens, respectively, interphotoreceptor binding protein (IRBP) and S-antigen (S-Ag). B10-A mice that received a single unilateral injection of Ad-vIL-10 in the retro-orbital sinus venosus performed 1 day before immunization with IRBP in the footpads showed high levels of circulating vIL-10 in their sera and a significant reduction in pathological ocular manifestations. Lower levels of IFN-γ and IL-2 were found in cellular supernatants from IRBP-stimulated splenic cells in these treated mice. The local effect on ocular disease of vIL-10 was neutralized completely by injection of a monoclonal anti-vIL-10 antibody, demonstrating the specificity of the treatment. To determine whether the transfer of the vIL-10 gene within the periocular tissues of the eye could prevent acute EAU, a subconjunctival injection of Ad-vIL-10 was performed in Lewis rats simultaneously with S-antigen in the footpads. This injection determined in situ vIL-10 expression with very low circulating vIL-10 and led to a significant reduction of EAU without affecting the systemic immune response. The present results suggest that Ad-mediated gene transfer resulting in systemic and local expression of vIL-10 provide a promising approach for the treatment of uveitis. PMID:12390308

  4. Evaluation of New Fluorescent Lipophosphoramidates for Gene Transfer and Biodistribution Studies after Systemic Administration

    PubMed Central

    Belmadi, Nawal; Berchel, Mathieu; Denis, Caroline; Berthe, Wilfried; Sibiril, Yann; Le Gall, Tony; Haelters, Jean-Pierre; Jaffres, Paul-Alain; Montier, Tristan

    2015-01-01

    The objective of lung gene therapy is to reach the respiratory epithelial cells in order to deliver a functional nucleic acid sequence. To improve the synthetic carrier’s efficacy, knowledge of their biodistribution and elimination pathways, as well as cellular barriers faced, depending on the administration route, is necessary. Indeed, the in vivo fate guides the adaptation of their chemical structure and formulation to increase their transfection capacity while maintaining their tolerance. With this goal, lipidic fluorescent probes were synthesized and formulated with cationic lipophosphoramidate KLN47 (KLN: Karine Le Ny). We found that such formulations present constant compaction properties and similar transfection results without inducing additional cytotoxicity. Next, biodistribution profiles of pegylated and unpegylated lipoplexes were compared after systemic injection in mice. Pegylation of complexes led to a prolonged circulation in the bloodstream, whereas their in vivo bioluminescent expression profiles were similar. Moreover, systemic administration of pegylated lipoplexes resulted in a transient liver toxicity. These results indicate that these new fluorescent compounds could be added into lipoplexes in small amounts without perturbing the transfection capacities of the formulations. Such additional properties allow exploration of the in vivo biodistribution profiles of synthetic carriers as well as the expression intensity of the reporter gene. PMID:26540038

  5. Genomic study of the Type IVC secretion system in Clostridium difficile: understanding C. difficile evolution via horizontal gene transfer.

    PubMed

    Zhang, Wen; Cheng, Ying; Du, Pengcheng; Zhang, Yuanyuan; Jia, Hongbing; Li, Xianping; Wang, Jing; Han, Na; Qiang, Yujun; Chen, Chen; Lu, Jinxing

    2017-01-01

    Clostridium difficile, the etiological agent of Clostridium difficile infection (CDI), is a gram-positive, spore-forming bacillus that is responsible for ∼20% of antibiotic-related cases of diarrhea and nearly all cases of pseudomembranous colitis. Previous data have shown that a substantial proportion (11%) of the C. difficile genome consists of mobile genetic elements, including seven conjugative transposons. However, the mechanism underlying the formation of a mosaic genome in C. difficile is unknown. The type-IV secretion system (T4SS) is the only secretion system known to transfer DNA segments among bacteria. We searched genome databases to identify a candidate T4SS in C. difficile that could transfer DNA among different C. difficile strains. All T4SS gene clusters in C. difficile are located within genomic islands (GIs), which have variable lengths and structures and are all conjugative transposons. During the horizontal-transfer process of T4SS GIs within the C. difficile population, the excision sites were altered, resulting in different short-tandem repeat sequences among the T4SS GIs, as well as different chromosomal insertion sites and additional regions in the GIs.

  6. Analysis of Porphyra membrane transporters demonstrates gene transfer among photosynthetic eukaryotes and numerous sodium-coupled transport systems.

    PubMed

    Chan, Cheong Xin; Zäuner, Simone; Wheeler, Glen; Grossman, Arthur R; Prochnik, Simon E; Blouin, Nicolas A; Zhuang, Yunyun; Benning, Christoph; Berg, Gry Mine; Yarish, Charles; Eriksen, Renée L; Klein, Anita S; Lin, Senjie; Levine, Ira; Brawley, Susan H; Bhattacharya, Debashish

    2012-04-01

    Membrane transporters play a central role in many cellular processes that rely on the movement of ions and organic molecules between the environment and the cell, and between cellular compartments. Transporters have been well characterized in plants and green algae, but little is known about transporters or their evolutionary histories in the red algae. Here we examined 482 expressed sequence tag contigs that encode putative membrane transporters in the economically important red seaweed Porphyra (Bangiophyceae, Rhodophyta). These contigs are part of a comprehensive transcriptome dataset from Porphyra umbilicalis and Porphyra purpurea. Using phylogenomics, we identified 30 trees that support the expected monophyly of red and green algae/plants (i.e. the Plantae hypothesis) and 19 expressed sequence tag contigs that show evidence of endosymbiotic/horizontal gene transfer involving stramenopiles. The majority (77%) of analyzed contigs encode transporters with unresolved phylogenies, demonstrating the difficulty in resolving the evolutionary history of genes. We observed molecular features of many sodium-coupled transport systems in marine algae, and the potential for coregulation of Porphyra transporter genes that are associated with fatty acid biosynthesis and intracellular lipid trafficking. Although both the tissue-specific and subcellular locations of the encoded proteins require further investigation, our study provides red algal gene candidates associated with transport functions and novel insights into the biology and evolution of these transporters.

  7. A Gene Transfer Agent and a Dynamic Repertoire of Secretion Systems Hold the Keys to the Explosive Radiation of the Emerging Pathogen Bartonella

    PubMed Central

    Guy, Lionel; Nystedt, Björn; Toft, Christina; Zaremba-Niedzwiedzka, Katarzyna; Berglund, Eva C.; Granberg, Fredrik; Näslund, Kristina; Eriksson, Ann-Sofie; Andersson, Siv G. E.

    2013-01-01

    Gene transfer agents (GTAs) randomly transfer short fragments of a bacterial genome. A novel putative GTA was recently discovered in the mouse-infecting bacterium Bartonella grahamii. Although GTAs are widespread in phylogenetically diverse bacteria, their role in evolution is largely unknown. Here, we present a comparative analysis of 16 Bartonella genomes ranging from 1.4 to 2.6 Mb in size, including six novel genomes from Bartonella isolated from a cow, two moose, two dogs, and a kangaroo. A phylogenetic tree inferred from 428 orthologous core genes indicates that the deadly human pathogen B. bacilliformis is related to the ruminant-adapted clade, rather than being the earliest diverging species in the genus as previously thought. A gene flux analysis identified 12 genes for a GTA and a phage-derived origin of replication as the most conserved innovations. These are located in a region of a few hundred kb that also contains 8 insertions of gene clusters for type III, IV, and V secretion systems, and genes for putatively secreted molecules such as cholera-like toxins. The phylogenies indicate a recent transfer of seven genes in the virB gene cluster for a type IV secretion system from a cat-adapted B. henselae to a dog-adapted B. vinsonii strain. We show that the B. henselae GTA is functional and can transfer genes in vitro. We suggest that the maintenance of the GTA is driven by selection to increase the likelihood of horizontal gene transfer and argue that this process is beneficial at the population level, by facilitating adaptive evolution of the host-adaptation systems and thereby expansion of the host range size. The process counters gene loss and forces all cells to contribute to the production of the GTA and the secreted molecules. The results advance our understanding of the role that GTAs play for the evolution of bacterial genomes. PMID:23555299

  8. A gene transfer agent and a dynamic repertoire of secretion systems hold the keys to the explosive radiation of the emerging pathogen Bartonella.

    PubMed

    Guy, Lionel; Nystedt, Björn; Toft, Christina; Zaremba-Niedzwiedzka, Katarzyna; Berglund, Eva C; Granberg, Fredrik; Näslund, Kristina; Eriksson, Ann-Sofie; Andersson, Siv G E

    2013-03-01

    Gene transfer agents (GTAs) randomly transfer short fragments of a bacterial genome. A novel putative GTA was recently discovered in the mouse-infecting bacterium Bartonella grahamii. Although GTAs are widespread in phylogenetically diverse bacteria, their role in evolution is largely unknown. Here, we present a comparative analysis of 16 Bartonella genomes ranging from 1.4 to 2.6 Mb in size, including six novel genomes from Bartonella isolated from a cow, two moose, two dogs, and a kangaroo. A phylogenetic tree inferred from 428 orthologous core genes indicates that the deadly human pathogen B. bacilliformis is related to the ruminant-adapted clade, rather than being the earliest diverging species in the genus as previously thought. A gene flux analysis identified 12 genes for a GTA and a phage-derived origin of replication as the most conserved innovations. These are located in a region of a few hundred kb that also contains 8 insertions of gene clusters for type III, IV, and V secretion systems, and genes for putatively secreted molecules such as cholera-like toxins. The phylogenies indicate a recent transfer of seven genes in the virB gene cluster for a type IV secretion system from a cat-adapted B. henselae to a dog-adapted B. vinsonii strain. We show that the B. henselae GTA is functional and can transfer genes in vitro. We suggest that the maintenance of the GTA is driven by selection to increase the likelihood of horizontal gene transfer and argue that this process is beneficial at the population level, by facilitating adaptive evolution of the host-adaptation systems and thereby expansion of the host range size. The process counters gene loss and forces all cells to contribute to the production of the GTA and the secreted molecules. The results advance our understanding of the role that GTAs play for the evolution of bacterial genomes.

  9. Heat transfer system

    DOEpatents

    Not Available

    1980-03-07

    A heat transfer system for a nuclear reactor is described. Heat transfer is accomplished within a sealed vapor chamber which is substantially evacuated prior to use. A heat transfer medium, which is liquid at the design operating temperatures, transfers heat from tubes interposed in the reactor primary loop to spaced tubes connected to a steam line for power generation purposes. Heat transfer is accomplished by a two-phase liquid-vapor-liquid process as used in heat pipes. Condensible gases are removed from the vapor chamber through a vertical extension in open communication with the chamber interior.

  10. Long-Term Systemic Myostatin Inhibition via Liver-Targeted Gene Transfer in Golden Retriever Muscular Dystrophy

    PubMed Central

    Sleeper, Meg M.; Forbes, Sean C.; Morine, Kevin J.; Reynolds, Caryn; Singletary, Gretchen E.; Trafny, Dennis; Pham, Jennifer; Bogan, Janet; Kornegay, Joe N.; Vandenborne, Krista; Walter, Glenn A.; Sweeney, H. Lee

    2011-01-01

    Abstract Duchenne muscular dystrophy (DMD) is a lethal, X-linked recessive disease affecting 1 in 3,500 newborn boys for which there is no effective treatment or cure. One novel strategy that has therapeutic potential for DMD is inhibition of myostatin, a negative regulator of skeletal muscle mass that may also promote fibrosis. Therefore, our goal in this study was to evaluate systemic myostatin inhibition in the golden retriever model of DMD (GRMD). GRMD canines underwent liver-directed gene transfer of a self-complementary adeno-associated virus type 8 vector designed to express a secreted dominant-negative myostatin peptide (n=4) and were compared with age-matched, untreated GRMD controls (n=3). Dogs were followed with serial magnetic resonance imaging (MRI) for 13 months to assess cross-sectional area and volume of skeletal muscle, then euthanized so that tissue could be harvested for morphological and histological analysis. We found that systemic myostatin inhibition resulted in increased muscle mass in GRMD dogs as assessed by MRI and confirmed at tissue harvest. We also found that hypertrophy of type IIA fibers was largely responsible for the increased muscle mass and that reductions in serum creatine kinase and muscle fibrosis were associated with long-term myostatin inhibition in GRMD. This is the first report describing the effects of long-term, systemic myostatin inhibition in a large-animal model of DMD, and we believe that the simple and effective nature of our liver-directed gene-transfer strategy makes it an ideal candidate for evaluation as a novel therapeutic approach for DMD patients. PMID:21787232

  11. Clinical Applications Involving CNS Gene Transfer

    PubMed Central

    Kantor, Boris; McCown, Thomas; Leone, Paola; Gray, Steven J.

    2015-01-01

    Diseases of the central nervous system (CNS) have traditionally been the most difficult to treat by traditional pharmacological methods, due mostly to the blood–brain barrier and the difficulties associated with repeated drug administration targeting the CNS. Viral vector gene transfer represents a way to permanently provide a therapeutic protein within the nervous system after a single administration, whether this be a gene replacement strategy for an inherited disorder or a disease-modifying protein for a disease such as Parkinson's. Gene therapy approaches for CNS disorders has evolved considerably over the last two decades. Although a breakthrough treatment has remained elusive, current strategies are now considerably safer and potentially much more effective. This chapter will explore the past, current, and future status of CNS gene therapy, focusing on clinical trials utilizing adeno-associated virus and lentiviral vectors. PMID:25311921

  12. Extensive beta-glucuronidase activity in murine central nervous system after adenovirus-mediated gene transfer to brain.

    PubMed

    Ghodsi, A; Stein, C; Derksen, T; Yang, G; Anderson, R D; Davidson, B L

    1998-11-01

    Mucopolysaccharidosis type VII (MPS VII), caused by beta-glucuronidase deficiency, is a classic lysosomal storage disease. In the central nervous system (CNS), there is widespread pathology with distention of vacuoles in neurons and glia. An approach to therapy for MPS VII would require extensive delivery of enzyme to the CNS and subsequent uptake by the affected cells. In this study we show that intrastriatal injection of recombinant adenovirus encoding beta-glucuronidase (Ad betagluc) to MPS VII or wild-type mice results in focal, intense beta-glucuronidase mRNA expression near the injection site. Further, histochemical staining for enzyme activity showed that beta-glucuronidase activity extended well beyond transduced cells. Activity was detected throughout the ipsilateral striatum as well as in the corpus callosum, ventricles, and bilateral neocortex. Similarly, after injection into the right lateral ventricle or cisterna magna, enzyme activity was present in the ependymal cells of the ventricles, in the subarachnoid spaces, and also in the underlying cortex (150-500 microm from ependyma). The distribution of enzyme was most extensive 21 days after gene transfer to normal mouse brain, with more than 50% of the hemisphere positive for beta-glucuronidase activity. Eighty-four days after adenovirus injection a substantial level of enzyme expression remained (>40% of hemisphere positive for beta-glucuronidase activity). Histological sections from striatum of beta-glucuronidase-deficient mice injected with Ad betagluc showed a marked reduction in the number of distended vacuoles in both neurons and glia, as compared with uninjected striatum. Importantly, correction was noted in both hemispheres. Our finding that a relatively small number of transduced cells produce enzyme that reaches a large proportion of the CNS has favorable implications in developing direct gene transfer therapies for lysosomal storage disorders.

  13. Fuel transfer system

    DOEpatents

    Townsend, H.E.; Barbanti, G.

    1994-03-01

    A nuclear fuel bundle fuel transfer system includes a transfer pool containing water at a level above a reactor core. A fuel transfer machine therein includes a carriage disposed in the transfer pool and under the water for transporting fuel bundles. The carriage is selectively movable through the water in the transfer pool and individual fuel bundles are carried vertically in the carriage. In a preferred embodiment, a first movable bridge is disposed over an upper pool containing the reactor core, and a second movable bridge is disposed over a fuel storage pool, with the transfer pool being disposed therebetween. A fuel bundle may be moved by the first bridge from the reactor core and loaded into the carriage which transports the fuel bundle to the second bridge which picks up the fuel bundle and carries it to the fuel storage pool. 6 figures.

  14. Fuel transfer system

    DOEpatents

    Townsend, Harold E.; Barbanti, Giancarlo

    1994-01-01

    A nuclear fuel bundle fuel transfer system includes a transfer pool containing water at a level above a reactor core. A fuel transfer machine therein includes a carriage disposed in the transfer pool and under the water for transporting fuel bundles. The carriage is selectively movable through the water in the transfer pool and individual fuel bundles are carried vertically in the carriage. In a preferred embodiment, a first movable bridge is disposed over an upper pool containing the reactor core, and a second movable bridge is disposed over a fuel storage pool, with the transfer pool being disposed therebetween. A fuel bundle may be moved by the first bridge from the reactor core and loaded into the carriage which transports the fuel bundle to the second bridge which picks up the fuel bundle and carries it to the fuel storage pool.

  15. Stable transformation of maize after gene transfer by electroporation.

    PubMed

    Fromm, M E; Taylor, L P; Walbot, V

    The graminaceous monocots, including the economically important cereals, seem to be refractory to infection by Agrobacterium tumefaciens, a natural gene transfer system that has been successfully exploited for transferring foreign genes into higher plants. Therefore, direct transfer techniques that are potentially applicable to all plant species have been developed using a few dicot and monocot species as model systems. One of these techniques, electroporation, uses electrical pulses of high field strength to permeabilize cell membranes reversibly so as to facilitate the transfer of DNA into cells. Electroporation-mediated gene transfer has resulted in stably transformed animal cells and transient gene expression in monocot and dicot plant cells. Here we report that electroporation-mediated DNA transfer of a chimaeric gene encoding neomycin phosphotransferase results in stably transformed maize cells that are resistant to kanamycin.

  16. Lateral gene transfer in the subsurface

    SciTech Connect

    Barkay, Tamar; Sobecky, Patricia

    2007-08-27

    Lateral gene transfer (LGT) is an important adaptive mechanism among prokaryotic organisms. This mechanism is particularly important for the response of microorganisms to changing environmental conditions because it facilitates the transfer of a large number of genes and their rapid expression. Together the transferred genes promote rapid genetic and metabolic changes that may enhance survival to newly established and sometimes hostile environmental conditions. The goal of our project was to examine if and how LGT enhances microbial adaptation to toxic heavy metals in subsurface environments that had been contaminated by mixed wastes due to activities associated with the production of nuclear energy and weapons. This task has been accomplished by dividing the project to several sub-tasks. Thus, we: (1) Determined the level of resistance of subsurface bacterial isolates to several toxic metals, all identified as pollutants of concern in subsurface environments; (2) Designed, tested, and applied, a molecular approach that determined whether metal resistance genes had evolved by LGT among subsurface bacteria; and (3) Developed a DNA hybridization array for the identification of broad host range plasmids and of metal resistance plasmids. The results are briefly summarized below with references to published papers and manuscripts in preparation where details about our research can be found. Additional information may be found in copies of our published manuscripts and conference proceedings, and our yearly reports that were submitted through the RIMS system.

  17. Keratinocyte gene therapy for systemic diseases. Circulating interleukin 10 released from gene-transferred keratinocytes inhibits contact hypersensitivity at distant areas of the skin.

    PubMed Central

    Meng, X; Sawamura, D; Tamai, K; Hanada, K; Ishida, H; Hashimoto, I

    1998-01-01

    This study has examined the systemic effects of a circulating gene product, human interleukin 10 (IL-10), released from transduced keratinocytes. IL-10 is an anti-inflammatory cytokine which has an inhibitory effect on contact hypersensitivity (CHS). An expression vector (phIL-10) was constructed for human IL-10 and was injected into the dorsal skin of hairless rats. Local expression of IL-10 mRNA and protein was detected by reverse-transcriptase polymerase chain reaction and immunohistochemical staining, respectively. Enzyme-linked immunosorbent assay showed that the amount of IL-10 in the local keratinocytes and in the circulation increased with the dose of phIL-10 transferred. To determine whether circulating IL-10 could inhibit the effector phase of CHS at a distant area of the skin, various doses of phIL-10 were injected into the dorsal skin of sensitized rats before challenge on the ears. Our results showed that the degree of swelling of the ears of phIL-10- treated rats was significantly lower than that in the negative control animals. These results suggest that IL-10 released from transduced keratinocytes can enter the bloodstream and cause biological effects at distant areas of the skin. This study demonstrates that it may be possible to treat systemic disease using keratinocyte gene therapy. PMID:9502789

  18. Horizontal gene transfer of chromosomal Type II toxin-antitoxin systems of Escherichia coli.

    PubMed

    Ramisetty, Bhaskar Chandra Mohan; Santhosh, Ramachandran Sarojini

    2016-02-01

    Type II toxin-antitoxin systems (TAs) are small autoregulated bicistronic operons that encode a toxin protein with the potential to inhibit metabolic processes and an antitoxin protein to neutralize the toxin. Most of the bacterial genomes encode multiple TAs. However, the diversity and accumulation of TAs on bacterial genomes and its physiological implications are highly debated. Here we provide evidence that Escherichia coli chromosomal TAs (encoding RNase toxins) are 'acquired' DNA likely originated from heterologous DNA and are the smallest known autoregulated operons with the potential for horizontal propagation. Sequence analyses revealed that integration of TAs into the bacterial genome is unique and contributes to variations in the coding and/or regulatory regions of flanking host genome sequences. Plasmids and genomes encoding identical TAs of natural isolates are mutually exclusive. Chromosomal TAs might play significant roles in the evolution and ecology of bacteria by contributing to host genome variation and by moderation of plasmid maintenance.

  19. Orbital Fluid Transfer System

    NASA Technical Reports Server (NTRS)

    Johnston, A. S., (Nick); Ryder, Mel; Tyler, Tony R.

    1998-01-01

    An automated fluid and power interface system needs to be developed for future space missions which require on orbit consumable replenishment. Current method of fluid transfer require manned vehicles and extravehicular activity. Currently the US does not have an automated capability for consumable transfer on-orbit. This technology would benefit both Space Station and long duration satellites. In order to provide this technology the Automated Fluid Interface System (AFIS) was developed. The AFIS project was an advanced development program aimed at developing a prototype satellite servicer for future space operations. This mechanism could transfer propellants, cryogens, fluids, gasses, electrical power, and communications from a tanker unit to the orbiting satellite. The development of this unit was a cooperative effort between Marshall Space Flight Center in Huntsville, Alabama, and Moog, Inc. in East Aurora, New York. An engineering model was built and underwent substantial development testing at Marshall Space Flight Center (MSFC). While the AFIS is not suitable for spaceflight, testing and evaluation of the AFIS provided significant experience which would be beneficial in building a flight unit. The lessons learned from testing the AFIS provided the foundation for the next generation fluid transfer mechanism, the Orbital Fluid Transfer System (OFTS). The OFTS project was a study contract with MSFC and Moog, Inc. The OFTS was designed for the International Space Station (ISS), but its flexible design could used for long duration satellite missions and other applications. The OFTS was designed to be used after docking. The primary function was to transfer bipropellants and high pressure gases. The other items addressed by this task included propellant storage, hardware integration, safety and control system issues. A new concept for high pressure couplings was also developed. The results of the AFIS testing provided an excellent basis for the OFTS design. The OFTS

  20. Targeting Radiotherapy to Cancer by Gene Transfer

    PubMed Central

    2003-01-01

    Targeted radionuclide therapy is an alternative method of radiation treatment which uses a tumor-seeking agent carrying a radioactive atom to deposits of tumor, wherever in the body they may be located. Recent experimental data signifies promise for the amalgamation of gene transfer with radionuclide targeting. This review encompasses aspects of the integration of gene manipulation and targeted radiotherapy, highlighting the possibilities of gene transfer to assist the targeting of cancer with low molecular weight radiopharmaceuticals. PMID:12721515

  1. Horizontal gene transfer in parasitic plants.

    PubMed

    Davis, Charles C; Xi, Zhenxiang

    2015-08-01

    Horizontal gene transfer (HGT) between species has been a major focus of plant evolutionary research during the past decade. Parasitic plants, which establish a direct connection with their hosts, have provided excellent examples of how these transfers are facilitated via the intimacy of this symbiosis. In particular, phylogenetic studies from diverse clades indicate that parasitic plants represent a rich system for studying this phenomenon. Here, HGT has been shown to be astonishingly high in the mitochondrial genome, and appreciable in the nuclear genome. Although explicit tests remain to be performed, some transgenes have been hypothesized to be functional in their recipient species, thus providing a new perspective on the evolution of novelty in parasitic plants.

  2. Lateral Gene Transfer from the Dead

    PubMed Central

    Szöllősi, Gergely J.; Tannier, Eric; Lartillot, Nicolas; Daubin, Vincent

    2013-01-01

    In phylogenetic studies, the evolution of molecular sequences is assumed to have taken place along the phylogeny traced by the ancestors of extant species. In the presence of lateral gene transfer, however, this may not be the case, because the species lineage from which a gene was transferred may have gone extinct or not have been sampled. Because it is not feasible to specify or reconstruct the complete phylogeny of all species, we must describe the evolution of genes outside the represented phylogeny by modeling the speciation dynamics that gave rise to the complete phylogeny. We demonstrate that if the number of sampled species is small compared with the total number of existing species, the overwhelming majority of gene transfers involve speciation to and evolution along extinct or unsampled lineages. We show that the evolution of genes along extinct or unsampled lineages can to good approximation be treated as those of independently evolving lineages described by a few global parameters. Using this result, we derive an algorithm to calculate the probability of a gene tree and recover the maximum-likelihood reconciliation given the phylogeny of the sampled species. Examining 473 near-universal gene families from 36 cyanobacteria, we find that nearly a third of transfer events (28%) appear to have topological signatures of evolution along extinct species, but only approximately 6% of transfers trace their ancestry to before the common ancestor of the sampled cyanobacteria. [Gene tree reconciliation; lateral gene transfer; macroevolution; phylogeny.] PMID:23355531

  3. Wireless power transfer system

    DOEpatents

    Wu, Hunter; Sealy, Kylee; Gilchrist, Aaron

    2016-02-23

    A system includes a first stage of an inductive power transfer system with an LCL load resonant converter with a switching section, an LCL tuning circuit, and a primary receiver pad. The IPT system includes a second stage with a secondary receiver pad, a secondary resonant circuit, a secondary rectification circuit, and a secondary decoupling converter. The secondary receiver pad connects to the secondary resonant circuit. The secondary resonant circuit connects to the secondary rectification circuit. The secondary rectification circuit connects to the secondary decoupling converter. The second stage connects to a load. The load includes an energy storage element. The second stage and load are located on a vehicle and the first stage is located at a fixed location. The primary receiver pad wirelessly transfers power to the secondary receiver pad across a gap when the vehicle positions the secondary receiver pad with respect to the primary receiver pad.

  4. Biased gene transfer in microbial evolution.

    PubMed

    Andam, Cheryl P; Gogarten, J Peter

    2011-06-13

    Horizontal gene transfer (HGT) is an important evolutionary process that allows the spread of innovations between distantly related organisms. We present evidence that prokaryotes (bacteria and archaea) are more likely to transfer genetic material with their close relatives than with distantly related lineages. This bias in transfer partners can create phylogenetic signals that are difficult to distinguish from the signal created through shared ancestry. Preferences for transfer partners can be revealed by studying the distribution patterns of divergent genes with identical functions. In many respects, these genes are similar to alleles in a population, except that they coexist only in higher taxonomic groupings and are acquired by a species through HGT. We also discuss the role of biased gene transfer in the formation of taxonomically recognizable natural groups in the tree or net of life.

  5. Creation and validation of a widely applicable multiple gene transfer vector system for stable transformation in plant.

    PubMed

    Sun, Quanxi; Liu, Jiang; Li, Yaxiao; Zhang, Qin; Shan, Shihua; Li, Xinzheng; Qi, Baoxiu

    2013-11-01

    Multiple gene transfer (MGT) technology has become a powerful tool for basic and applied plant biology research in recent years. Despite some notable successes in obtaining plant lines harbouring multiple transgenes, these methods are still generally unwieldy and costly. We report here a straightforward and cost effective strategy, utilizing commonly available restriction enzymes for the transfer of multiple genes into plants, hence greatly widening the accessibility of MGT. This methodology exploits the specific 'nested' arrangement of a pair of isocaudomer restriction enzymes (for example XbaI-AvrII-XbaI) so that through the alternate use of these two enzymes in a reiterative fashion multiple genes/constructs (up to five in this study) could be 'stacked' together with ease. In a proof-of-concept experiment, we constructed a plant transformation vector containing three reporter gene expression cassettes flanked by two matrix attachment region sequences. The expression of all three genes was confirmed in transgenic Arabidopsis thaliana. The usefulness of this technology was further validated by the construction of a plant transformation vector containing five transgenes for the production of eicosapentaenoic acid (EPA, C20∆⁵,⁸,¹¹,¹⁴,¹⁷), a polyunsaturated essential fatty acid found in fish oils that is beneficial for health. In addition, we constructed four more vectors, incorporating one seed specific and three promoters conferring constitutive expression. These expression cassettes are flanked by a different isocaudomer pair (AvrII-SpeI-AvrII) and four other unique restriction sites, allowing the exchange of promoters and terminators of choice.

  6. CURRENT TRANSFER SYSTEMS

    DOEpatents

    Watt, D.A.

    1956-07-01

    A current transfer system is described for transferring current between a rotating member and a co-axial stationary member. The particular area of application for the invention is in connection with homopolar generators where a low voltage and high current are generated. The current tramsfer system of the invention comprises a rotor member and a co-axial stator member wherein one of the members is shaped to provide a circumferential surface concave in section and the other member is shaped to have a peripheral portion in close proximity to the surface, whereby a liquid metal can be stably supported between the two members when they are moving relative to one another to establish an electrical conducting path between the members.

  7. Detecting Highways of Horizontal Gene Transfer

    NASA Astrophysics Data System (ADS)

    Bansal, Mukul S.; Gogarten, J. Peter; Shamir, Ron

    In a horizontal gene transfer (HGT) event a gene is transferred between two species that do not share an ancestor-descendant relationship. Typically, no more than a few genes are horizontally transferred between any two species. However, several studies identified pairs of species between which many different genes were horizontally transferred. Such a pair is said to be linked by a highway of gene sharing. We present a method for inferring such highways. Our method is based on the fact that the evolutionary histories of horizontally transferred genes disagree with the corresponding species phylogeny. Specifically, given a set of gene trees and a trusted rooted species tree, each gene tree is first decomposed into its constituent quartet trees and the quartets that are inconsistent with the species tree are identified. Our method finds a pair of species such that a highway between them explains the largest (normalized) fraction of inconsistent quartets. For a problem on n species, our method requires O(n 4) time, which is optimal with respect to the quartets input size. An application of our method to a dataset of 1128 genes from 11 cyanobacterial species, as well as to simulated datasets, illustrates the efficacy of our method.

  8. Detecting highways of horizontal gene transfer.

    PubMed

    Bansal, Mukul S; Banay, Guy; Gogarten, J Peter; Shamir, Ron

    2011-09-01

    In a horizontal gene transfer (HGT) event, a gene is transferred between two species that do not have an ancestor-descendant relationship. Typically, no more than a few genes are horizontally transferred between any two species. However, several studies identified pairs of species between which many different genes were horizontally transferred. Such a pair is said to be linked by a highway of gene sharing. We present a method for inferring such highways. Our method is based on the fact that the evolutionary histories of horizontally transferred genes disagree with the corresponding species phylogeny. Specifically, given a set of gene trees and a trusted rooted species tree, each gene tree is first decomposed into its constituent quartet trees and the quartets that are inconsistent with the species tree are identified. Our method finds a pair of species such that a highway between them explains the largest (normalized) fraction of inconsistent quartets. For a problem on n species and m input quartet trees, we give an efficient O(m + n(2))-time algorithm for detecting highways, which is optimal with respect to the quartets input size. An application of our method to a dataset of 1128 genes from 11 cyanobacterial species, as well as to simulated datasets, illustrates the efficacy of our method.

  9. Thermal flux transfer system

    NASA Technical Reports Server (NTRS)

    Freggens, R. A. (Inventor)

    1973-01-01

    A thermal flux transfer system for use in maintaining the thrust chamber of an operative reaction motor at given temperatures is described. The system is characterized by an hermetically sealed chamber surrounding a thrust chamber to be cooled, with a plurality of parallel, longitudinally spaced, disk-shaped wick members formed of a metallic mesh and employed in delivering a working fluid, in its liquid state, radially toward the thrust chamber and delivering the working fluid, in its vapor state, away from the nozzle for effecting a cooling of the nozzle, in accordance with known principles of an operating heat pipe.

  10. Gene Transfers Between Distantly Related Organisms

    NASA Technical Reports Server (NTRS)

    Doolittle, Russell F.

    2003-01-01

    With the completion of numerous microbial genome sequences, reports of individual gene transfers between distantly related prokaryotes have become commonplace. On the other hand, transfers between prokaryotes and eukaryotes still excite the imagination. Many of these claims may be premature, but some are certainly valid. In this chapter, the kinds of supporting data needed to propose transfers between distantly related organisms and cite some interesting examples are considered.

  11. Antibody Gene Transfer for HIV Immunoprophylaxis

    PubMed Central

    Balazs, Alejandro B.; West, Anthony P.

    2015-01-01

    Antibody gene transfer, which involves the delivery of genes that encode potent, broadly neutralizing anti-HIV antibodies, is a promising new strategy to prevent HIV infection. A satellite symposium at the AIDS Vaccine 2012 conference brought together many of the groups working in this field. PMID:23238748

  12. Antibody gene transfer for HIV immunoprophylaxis.

    PubMed

    Balazs, Alejandro B; West, Anthony P

    2013-01-01

    Antibody gene transfer, which involves the delivery of genes that encode potent, broadly neutralizing antibodies to human immunodeficiency virus (HIV), is a promising new strategy for preventing HIV infection. A satellite symposium at the AIDS Vaccine 2012 conference brought together many of the groups working in this field.

  13. Occurrence and expression of gene transfer agent genes in marine bacterioplankton.

    PubMed

    Biers, Erin J; Wang, Kui; Pennington, Catherine; Belas, Robert; Chen, Feng; Moran, Mary Ann

    2008-05-01

    Genes with homology to the transduction-like gene transfer agent (GTA) were observed in genome sequences of three cultured members of the marine Roseobacter clade. A broader search for homologs for this host-controlled virus-like gene transfer system identified likely GTA systems in cultured Alphaproteobacteria, and particularly in marine bacterioplankton representatives. Expression of GTA genes and extracellular release of GTA particles ( approximately 50 to 70 nm) was demonstrated experimentally for the Roseobacter clade member Silicibacter pomeroyi DSS-3, and intraspecific gene transfer was documented. GTA homologs are surprisingly infrequent in marine metagenomic sequence data, however, and the role of this lateral gene transfer mechanism in ocean bacterioplankton communities remains unclear.

  14. Gene transfer in inner ear cells: a challenging race.

    PubMed

    Sacheli, R; Delacroix, L; Vandenackerveken, P; Nguyen, L; Malgrange, B

    2013-03-01

    Recent advances in human genomics led to the identification of numerous defective genes causing deafness, which represent novel putative therapeutic targets. Future gene-based treatment of deafness resulting from genetic or acquired sensorineural hearing loss may include strategies ranging from gene therapy to antisense delivery. For successful development of gene therapies, a minimal requirement involves the engineering of appropriate gene carrier systems. Transfer of exogenous genetic material into the mammalian inner ear using viral or non-viral vectors has been characterized over the last decade. The nature of inner ear cells targeted, as well as the transgene expression level and duration, are highly dependent on the vector type, the route of administration and the strength of the promoter driving expression. This review summarizes and discusses recent advances in inner ear gene-transfer technologies aimed at examining gene function or identifying new treatment for inner ear disorders.

  15. Gene transfer as a future therapy for rheumatoid arthritis.

    PubMed

    Müller-Ladner, Ulf; Pap, Thomas; Gay, Renate E; Gay, Steffen

    2003-07-01

    Inhibiting key pathogenic processes within the rheumatoid synovium is a most attractive goal to achieve, and the number of potential intra- and extracellular pathways operative in rheumatoid arthritis (RA) that could be used for a gene therapy strategy is increasing continuously. Gene transfer or gene therapy might also be one of the approaches to solve the problem of long-term expression of therapeutic genes, in order to replace the frequent application of recombinant proteins, in the future. However, at present, gene therapy has not reached a realistic clinical stage, which is mainly due to severe side effects in humans, the complexity of RA pathophysiology and the current state of available gene transfer techniques. On the other hand, novel gene delivery systems are not restricted to vectors or certain types of cells, as mobile cells including macrophages, dendritic cells, lymphocytes and multipotent stem cells can also be used as smart gene transfer vehicles. Moreover, the observation in animal models that application of viral vectors into a joint can exert additional therapeutic effects in nearby joints might also facilitate the transfer from animal to human gene therapy. Future strategies will also examine the potential of novel long-term expression vectors such as lentiviruses and cytomegalovirus (CMV)-based viruses as a basis for future clinical trials in RA.

  16. Gene transfer mediated by alpha2-macroglobulin.

    PubMed Central

    Schneider, H; Huse, K; Birkenmeier, G; Otto, A; Scholz, G H

    1996-01-01

    alpha2-Macroglobulin covalently linked to poly(L)-lysine can be used as a vehicle for receptor-mediated gene transfer. This modified alpha2-macroglobulin maintains its ability to bind to the alpha2-macroglobulin receptor, and was shown to introduce a luciferase reporter gene plasmid into HepG2 human hepatoma cells in vitro. The alpha2-macroglobulin receptor is a very large and multifunctional cell surface receptor, whose rapid and efficient internalization rate makes it attractive for gene therapy, e.g. for hepatic gene targeting via injection into the portal vein. PMID:8871570

  17. Molecular Transfer of Nematode Resistance Genes

    PubMed Central

    Williamson, V. M.; Ho, J.-Y.; Ma, H. M.

    1992-01-01

    Recombinant DNA techniques have been used to introduce agronomically valuable traits, including resistance to viruses, herbicides, and insects, into crop plants. Introduction of these genes into plants frequently involves Agrobacterium-mediated gene transfer. The potential exists for applying this technology to nematode control by introducing genes conferring resistance to nematodes. Transferred genes could include those encoding products detrimental to nematode development or reproduction as well as cloned host resistance genes. Host genes that confer resistance to cyst or root-knot nematode species have been identified in many plants. The best characterized is Mi, a gene that confers resistance to root-knot nematodes in tomato. A map-based cloning approach is being used to isolate the gene. For development of a detailed map of the region of the genome surrounding Mi, DNA markers genetically linked to Mi have been identified and analyzed in tomato lines that have undergone a recombination event near Mi. The molecular map will be used to identify DNA corresponding to Mi. We estimate that a clone of Mi will be obtained in 2-5 years. An exciting prospect is that introduction of this gene will confer resistance in plant species without currently available sources of resistance. PMID:19282989

  18. Viral Vectors for in Vivo Gene Transfer

    NASA Astrophysics Data System (ADS)

    Thévenot, E.; Dufour, N.; Déglon, N.

    The transfer of DNA into the nucleus of a eukaryotic cell (gene transfer) is a central theme of modern biology. The transfer is said to be somatic when it refers to non-germline organs of a developed individual, and germline when it concerns gametes or the fertilised egg of an animal, with the aim of transmitting the relevant genetic modification to its descendents [1]. The efficient introduction of genetic material into a somatic or germline cell and the control of its expression over time have led to major advances in understanding how genes work in vivo, i.e., in living organisms (functional genomics), but also to the development of innovative therapeutic methods (gene therapy). The efficiency of gene transfer is conditioned by the vehicle used, called the vector. Desirable features for a vector are as follows: Easy to produce high titer stocks of the vector in a reproducible way. Absence of toxicity related to transduction (transfer of genetic material into the target cell, and its expression there) and no immune reaction of the organism against the vector and/or therapeutic protein. Stability in the expression of the relevant gene over time, and the possibility of regulation, e.g., to control expression of the therapeutic protein on the physiological level, or to end expression at the end of treatment. Transduction of quiescent cells should be as efficient as transduction of dividing cells. Vectors currently used fall into two categories: non-viral and viral vectors. In non-viral vectors, the DNA is complexed with polymers, lipids, or cationic detergents (described in Chap. 3). These vectors have a low risk of toxicity and immune reaction. However, they are less efficient in vivo than viral vectors when it comes to the number of cells transduced and long-term transgene expression. (Naked DNA transfer or electroporation is rather inefficient in the organism. This type of gene transfer will not be discussed here, and the interested reader is referred to the

  19. Muscle as a target for supplementary factor IX gene transfer.

    PubMed

    Hoffman, Brad E; Dobrzynski, Eric; Wang, Lixin; Hirao, Lauren; Mingozzi, Federico; Cao, Ou; Herzog, Roland W

    2007-07-01

    Immune responses to the factor IX (F.IX) transgene product are a concern in gene therapy for the X-linked bleeding disorder hemophilia B. The risk for such responses is determined by several factors, including the vector, target tissue, and others. Previously, we have demonstrated that hepatic gene transfer with adeno-associated viral (AAV) vectors can induce F.IX-specific immune tolerance. Muscle-derived F.IX expression, however, is limited by a local immune response. Here, skeletal muscle was investigated as a target for supplemental gene transfer. Given the low invasiveness of intramuscular injections, this route would be ideal for secondary gene transfer, thereby boosting levels of transgene expression. However, this is feasible only if immune tolerance established by compartmentalization of expression to the liver extends to other sites. Immune tolerance to human F.IX established by prior hepatic AAV-2 gene transfer was maintained after subsequent injection of AAV-1 or adenoviral vector into skeletal muscle, and tolerized mice failed to form antibodies or an interferon (IFN)-gamma(+) T cell response to human F.IX. A sustained increase in systemic transgene expression was obtained for AAV-1, whereas an increase after adenoviral gene transfer was transient. A CD8(+) T cell response specifically against adenovirus-transduced fibers was observed, suggesting that cytotoxic T cell responses against viral antigens were sufficient to eliminate expression in muscle. In summary, the data demonstrate that supplemental F.IX gene transfer to skeletal muscle does not break tolerance achieved by liver-derived expression. The approach is efficacious, if the vector for muscle gene transfer does not express immunogenic viral proteins.

  20. Heat Transfer Parametric System Identification

    DTIC Science & Technology

    1993-06-01

    Transfer Parametric System Identification 6. AUTHOR(S Parker, Gregory K. 7. PERFORMING ORGANIZATION NAME(S) AND AOORESS(ES) 8. PERFORMING ORGANIZATION...distribution is unlimited. Heat Transfer Parametric System Identification by Gregory K. Parker Lieutenant, United States Navy BS., DeVry Institute of...Modeling Concept ........ ........... 3 2. Lumped Parameter Approach ...... ......... 4 3. Parametric System Identification ....... 4 B. BASIC MODELING

  1. Overexpression of lipid transfer protein (LTP) genes enhances resistance to plant pathogens and LTP functions in long-distance systemic signaling in tobacco.

    PubMed

    Sarowar, Sujon; Kim, Young Jin; Kim, Ki Deok; Hwang, Byung Kook; Ok, Sung Han; Shin, Jeong Sheop

    2009-03-01

    The lipid signal is essential for the activation of plant defense responses, but downstream components of the signaling pathway are still poorly defined. To investigate the biological functions of pepper lipid transfer protein (LTP), we carried out virus-induced gene silencing (VIGS) in pepper, constitutive expression of CALTPs and grafting experiments in the tobacco plant. Suppression of endogenous CALTPI and CALTPII by VIGS, respectively, resulted in enhanced susceptibility to Xanthomonas campestris pv. vescatoria and pepper mosaic mottle virus in pepper. On the other hand, the constitutive expression of CALTPI and CALTPII genes in tobacco plants showed enhanced resistance to oomycete pathogen, Phytophthora nicotianae and bacterial pathogen, Pseudomonas syringae pv. tabaci. Enhanced resistance is found to be associated with the enhanced CALTP transcript levels in the independent transgenic CALTPI or II tobacco lines. Induced resistance responses in grafted scion leaves revealed that LTP plays a role in long-distance systemic signaling in plants.

  2. Vector-mediated antibody gene transfer for infectious diseases.

    PubMed

    Schnepp, Bruce C; Johnson, Philip R

    2015-01-01

    This chapter discusses the emerging field of vector-mediated antibody gene transfer as an alternative vaccine for infectious disease, with a specific focus on HIV. However, this methodology need not be confined to HIV-1; the general strategy of vector-mediated antibody gene transfer can be applied to other difficult vaccine targets like hepatitis C virus, malaria, respiratory syncytial virus, and tuberculosis. This approach is an improvement over classical passive immunization strategies that administer antibody proteins to the host to provide protection from infection. With vector-mediated gene transfer, the antibody gene is delivered to the host, via a recombinant adeno-associated virus (rAAV) vector; this in turn results in long-term endogenous antibody expression from the injected muscle that confers protective immunity. Vector-mediated antibody gene transfer can rapidly move existing, potent broadly cross-neutralizing HIV-1-specific antibodies into the clinic. The gene transfer products demonstrate a potency and breadth identical to the original product. This strategy eliminates the need for immunogen design and interaction with the adaptive immune system to generate protection, a strategy that so far has shown limited promise.

  3. Emerging role of regulatory T cells in gene transfer.

    PubMed

    Cao, Ou; Furlan-Freguia, Christian; Arruda, Valder R; Herzog, Roland W

    2007-10-01

    Induction and maintenance of immune tolerance to therapeutic transgene products are key requirements for successful gene replacement therapies. Gene transfer may also be used to specifically induce immune tolerance and thereby augment other types of therapies. Similarly, gene therapies for treatment of autoimmune diseases are being developed in order to restore tolerance to self-antigens. Regulatory T cells have emerged as key players in many aspects of immune tolerance, and a rapidly increasing body of work documents induction and/or activation of regulatory T cells by gene transfer. Regulatory T cells may suppress antibody formation and cytotoxic T cell responses and may be critical for immune tolerance to therapeutic proteins. In this regard, CD4(+)CD25(+) regulatory T cells have been identified as important components of tolerance in several gene transfer protocols, including hepatic in vivo gene transfer. Augmentation of regulatory T cell responses should be a promising new tool to achieve tolerance and avoid immune-mediated rejection of gene therapy. During the past decade, it has become obvious that immune regulation is an important and integral component of tolerance to self-antigens and of many forms of induced tolerance. Gene therapy can only be successful if the immune system does not reject the therapeutic transgene product. Recent studies provide a rapidly growing body of evidence that regulatory T cells (T(reg)) are involved and often play a crucial role in tolerance to proteins expressed by means of gene transfer. This review seeks to provide an overview of these data and their implications for gene therapy.

  4. Heat transfer in aeropropulsion systems

    NASA Technical Reports Server (NTRS)

    Simoneau, R. J.

    1985-01-01

    Aeropropulsion heat transfer is reviewed. A research methodology based on a growing synergism between computations and experiments is examined. The aeropropulsion heat transfer arena is identified as high Reynolds number forced convection in a highly disturbed environment subject to strong gradients, body forces, abrupt geometry changes and high three dimensionality - all in an unsteady flow field. Numerous examples based on heat transfer to the aircraft gas turbine blade are presented to illustrate the types of heat transfer problems which are generic to aeropropulsion systems. The research focus of the near future in aeropropulsion heat transfer is projected.

  5. Heat transfer in aeropropulsion systems

    NASA Astrophysics Data System (ADS)

    Simoneau, R. J.

    1985-07-01

    Aeropropulsion heat transfer is reviewed. A research methodology based on a growing synergism between computations and experiments is examined. The aeropropulsion heat transfer arena is identified as high Reynolds number forced convection in a highly disturbed environment subject to strong gradients, body forces, abrupt geometry changes and high three dimensionality - all in an unsteady flow field. Numerous examples based on heat transfer to the aircraft gas turbine blade are presented to illustrate the types of heat transfer problems which are generic to aeropropulsion systems. The research focus of the near future in aeropropulsion heat transfer is projected.

  6. Unsupervised Learning in Detection of Gene Transfer

    PubMed Central

    Hamel, L.; Nahar, N.; Poptsova, M. S.; Zhaxybayeva, O.; Gogarten, J. P.

    2008-01-01

    The tree representation as a model for organismal evolution has been in use since before Darwin. However, with the recent unprecedented access to biomolecular data, it has been discovered that, especially in the microbial world, individual genes making up the genome of an organism give rise to different and sometimes conflicting evolutionary tree topologies. This discovery calls into question the notion of a single evolutionary tree for an organism and gives rise to the notion of an evolutionary consensus tree based on the evolutionary patterns of the majority of genes in a genome embedded in a network of gene histories. Here, we discuss an approach to the analysis of genomic data of multiple genomes using bipartition spectral analysis and unsupervised learning. An interesting observation is that genes within genomes that have evolutionary tree topologies, which are in substantial conflict with the evolutionary consensus tree of an organism, point to possible horizontal gene transfer events which often delineate significant evolutionary events. PMID:18509479

  7. Viral vectors for gene transfer: current status of gene therapeutics.

    PubMed

    Heilbronn, Regine; Weger, Stefan

    2010-01-01

    Gene therapy for the correction of inherited or acquired disease has gained increasing importance in recent years. Successful treatment of children suffering from severe combined immunodeficiency (SCID) was achieved using retrovirus vectors for gene transfer. Encouraging improvements of vision were reported in a genetic eye disorder (LCA) leading to early childhood blindness. Adeno-associated virus (AAV) vectors were used for gene transfer in these trials. This chapter gives an overview of the design and delivery of viral vectors for the transport of a therapeutic gene into a target cell or tissue. The construction and production of retrovirus, lentivirus, and AAV vectors are covered. The focus is on production methods suitable for biopharmaceutical upscaling and for downstream processing. Quality control measures and biological safety considerations for the use of vectors in clinical trials are discussed.

  8. Perinatal Gene Transfer to the Liver

    PubMed Central

    McKay, Tristan R; Rahim, Ahad A; Buckley, Suzanne M.K; Ward, Natalie J; Chan, Jerry K.Y; Howe, Steven J; Waddington, Simon N

    2011-01-01

    The liver acts as a host to many functions hence raising the possibility that any one may be compromised by a single gene defect. Inherited or de novo mutations in these genes may result in relatively mild diseases or be so devastating that death within the first weeks or months of life is inevitable. Some diseases can be managed using conventional medicines whereas others are, as yet, untreatable. In this review we consider the application of early intervention gene therapy in neonatal and fetal preclinical studies. We appraise the tools of this technology, including lentivirus, adenovirus and adeno-associated virus (AAV)-based vectors. We highlight the application of these for a range of diseases including hemophilia, urea cycle disorders such as ornithine transcarbamylase deficiency, organic acidemias, lysosomal storage diseases including mucopolysaccharidoses, glycogen storage diseases and bile metabolism. We conclude by assessing the advantages and disadvantages associated with fetal and neonatal liver gene transfer. PMID:21774770

  9. Adenovirus-mediated gene transfer and expression of human beta-glucuronidase gene in the liver, spleen, and central nervous system in mucopolysaccharidosis type VII mice.

    PubMed

    Ohashi, T; Watabe, K; Uehara, K; Sly, W S; Vogler, C; Eto, Y

    1997-02-18

    Mucopolysaccharidosis type VII (Sly syndrome) is a lysosomal storage disease caused by inherited deficiency of the lysosomal enzyme beta-glucuronidase. A murine model of this disorder has been well characterized and used to study a number of forms of experimental therapies, including gene therapy. We produced recombinant adenovirus that expresses human beta-glucuronidase and administered this recombinant adenovirus to beta-glucuronidase-deficient mice intravenously. The beta-glucuronidase activities in liver and spleen were elevated to 40% and 20%, respectively, of the heterozygote enzymatic level at day 16. Expression persisted for at least 35 days. Pathological abnormalities of these tissues were also improved, and the elevated levels of urinary glycosaminoglycans were reduced in treated mice. However, the beta-glucuronidase activity in kidney and brain was not significantly increased. After administration of the recombinant adenovirus directly into the lateral ventricles of mutant mice, the beta-glucuronidase activity in crude brain homogenates increased to 30% of heterozygote activity. Histochemical demonstration of beta-glucuronidase activity in brain revealed that the enzymatic activity was mainly in ependymal cells and choroid. However, in some regions, the adenovirus-mediated gene expression was also evident in brain parenchyma associated with vessels and in the meninges. These results suggest that adenovirus-mediated gene delivery might improve the central nervous system pathology of mucopolysaccharidosis in addition to correcting visceral pathology.

  10. Sodium heat transfer system modeling

    NASA Astrophysics Data System (ADS)

    Baker, A. F.; Fewell, M. E.

    1983-11-01

    The sodium heat transfer system of the international energy agency (IEA) small solar power systems (SSPS) central receiver system (CRS), which includes the heliostat field, receiver, hot and cold storage vessels, and sodium/water steam generator was modeled. The computer code SOLTES (simulator of large thermal energy systems), was used to model this system. The results from SOLTES are compared to measured data.

  11. CANISTER TRANSFER SYSTEM DESCRIPTION DOCUMENT

    SciTech Connect

    B. Gorpani

    2000-06-23

    The Canister Transfer System receives transportation casks containing large and small disposable canisters, unloads the canisters from the casks, stores the canisters as required, loads them into disposal containers (DCs), and prepares the empty casks for re-shipment. Cask unloading begins with cask inspection, sampling, and lid bolt removal operations. The cask lids are removed and the canisters are unloaded. Small canisters are loaded directly into a DC, or are stored until enough canisters are available to fill a DC. Large canisters are loaded directly into a DC. Transportation casks and related components are decontaminated as required, and empty casks are prepared for re-shipment. One independent, remotely operated canister transfer line is provided in the Waste Handling Building System. The canister transfer line consists of a Cask Transport System, Cask Preparation System, Canister Handling System, Disposal Container Transport System, an off-normal canister handling cell with a transfer tunnel connecting the two cells, and Control and Tracking System. The Canister Transfer System operating sequence begins with moving transportation casks to the cask preparation area with the Cask Transport System. The Cask Preparation System prepares the cask for unloading and consists of cask preparation manipulator, cask inspection and sampling equipment, and decontamination equipment. The Canister Handling System unloads the canister(s) and places them into a DC. Handling equipment consists of a bridge crane hoist, DC loading manipulator, lifting fixtures, and small canister staging racks. Once the cask has been unloaded, the Cask Preparation System decontaminates the cask exterior and returns it to the Carrier/Cask Handling System via the Cask Transport System. After the DC is fully loaded, the Disposal Container Transport System moves the DC to the Disposal Container Handling System for welding. To handle off-normal canisters, a separate off-normal canister handling

  12. Canister Transfer System Description Document

    SciTech Connect

    2000-10-12

    The Canister Transfer System receives transportation casks containing large and small disposable canisters, unloads the canisters from the casks, stores the canisters as required, loads them into disposal containers (DCs), and prepares the empty casks for re-shipment. Cask unloading begins with cask inspection, sampling, and lid bolt removal operations. The cask lids are removed and the canisters are unloaded. Small canisters are loaded directly into a DC, or are stored until enough canisters are available to fill a DC. Large canisters are loaded directly into a DC. Transportation casks and related components are decontaminated as required, and empty casks are prepared for re-shipment. One independent, remotely operated canister transfer line is provided in the Waste Handling Building System. The canister transfer line consists of a Cask Transport System, Cask Preparation System, Canister Handling System, Disposal Container Transport System, an off-normal canister handling cell with a transfer tunnel connecting the two cells, and Control and Tracking System. The Canister Transfer System operating sequence begins with moving transportation casks to the cask preparation area with the Cask Transport System. The Cask Preparation System prepares the cask for unloading and consists of cask preparation manipulator, cask inspection and sampling equipment, and decontamination equipment. The Canister Handling System unloads the canister(s) and places them into a DC. Handling equipment consists of a bridge crane/hoist, DC loading manipulator, lifting fixtures, and small canister staging racks. Once the cask has been unloaded, the Cask Preparation System decontaminates the cask exterior and returns it to the Carrier/Cask Handling System via the Cask Transport System. After the DC is fully loaded, the Disposal Container Transport System moves the DC to the Disposal Container Handling System for welding. To handle off-normal canisters, a separate off-normal canister handling

  13. Systemic AAV9 gene transfer in adult GM1 gangliosidosis mice reduces lysosomal storage in CNS and extends lifespan

    PubMed Central

    Weismann, Cara M.; Ferreira, Jennifer; Keeler, Allison M.; Su, Qin; Qui, Linghua; Shaffer, Scott A.; Xu, Zuoshang; Gao, Guangping; Sena-Esteves, Miguel

    2015-01-01

    GM1 gangliosidosis (GM1) is an autosomal recessive lysosomal storage disease where GLB1 gene mutations result in a reduction or absence of lysosomal acid β-galactosidase (βgal) activity. βgal deficiency leads to accumulation of GM1-ganglioside in the central nervous system (CNS). GM1 is characterized by progressive neurological decline resulting in generalized paralysis, extreme emaciation and death. In this study, we assessed the therapeutic efficacy of an adeno-associated virus (AAV) 9-mβgal vector infused systemically in adult GM1 mice (βGal−/−) at 1 × 1011 or 3 × 1011 vector genomes (vg). Biochemical analysis of AAV9-treated GM1 mice showed high βGal activity in liver and serum. Moderate βGal levels throughout CNS resulted in a 36–76% reduction in GM1-ganglioside content in the brain and 75–86% in the spinal cord. Histological analyses of the CNS of animals treated with 3 × 1011 vg dose revealed increased presence of βgal and clearance of lysosomal storage throughout cortex, hippocampus, brainstem and spinal cord. Storage reduction in these regions was accompanied by a marked decrease in astrogliosis. AAV9 treatment resulted in improved performance in multiple tests of motor function and behavior. Also the majority of GM1 mice in the 3 × 1011 vg cohort retained ambulation and rearing despite reaching the humane endpoint due to weight loss. Importantly, the median survival of AAV9 treatment groups (316–576 days) was significantly increased over controls (250–264 days). This study shows that moderate widespread expression of βgal in the CNS of GM1 gangliosidosis mice is sufficient to achieve significant biochemical impact with phenotypic amelioration and extension in lifespan. PMID:25964428

  14. Systemic AAV9 gene transfer in adult GM1 gangliosidosis mice reduces lysosomal storage in CNS and extends lifespan.

    PubMed

    Weismann, Cara M; Ferreira, Jennifer; Keeler, Allison M; Su, Qin; Qui, Linghua; Shaffer, Scott A; Xu, Zuoshang; Gao, Guangping; Sena-Esteves, Miguel

    2015-08-01

    GM1 gangliosidosis (GM1) is an autosomal recessive lysosomal storage disease where GLB1 gene mutations result in a reduction or absence of lysosomal acid β-galactosidase (βgal) activity. βgal deficiency leads to accumulation of GM1-ganglioside in the central nervous system (CNS). GM1 is characterized by progressive neurological decline resulting in generalized paralysis, extreme emaciation and death. In this study, we assessed the therapeutic efficacy of an adeno-associated virus (AAV) 9-mβgal vector infused systemically in adult GM1 mice (βGal(-/-)) at 1 × 10(11) or 3 × 10(11) vector genomes (vg). Biochemical analysis of AAV9-treated GM1 mice showed high βGal activity in liver and serum. Moderate βGal levels throughout CNS resulted in a 36-76% reduction in GM1-ganglioside content in the brain and 75-86% in the spinal cord. Histological analyses of the CNS of animals treated with 3 × 10(11) vg dose revealed increased presence of βgal and clearance of lysosomal storage throughout cortex, hippocampus, brainstem and spinal cord. Storage reduction in these regions was accompanied by a marked decrease in astrogliosis. AAV9 treatment resulted in improved performance in multiple tests of motor function and behavior. Also the majority of GM1 mice in the 3 × 10(11) vg cohort retained ambulation and rearing despite reaching the humane endpoint due to weight loss. Importantly, the median survival of AAV9 treatment groups (316-576 days) was significantly increased over controls (250-264 days). This study shows that moderate widespread expression of βgal in the CNS of GM1 gangliosidosis mice is sufficient to achieve significant biochemical impact with phenotypic amelioration and extension in lifespan.

  15. Gene Transfer in Mycobacterium tuberculosis: Shuttle Phasmids to Enlightenment

    PubMed Central

    JACOBS, WILLIAM R.

    2016-01-01

    Infectious diseases have plagued humankind throughout history and have posed serious public health problems. Yet vaccines have eradicated smallpox and antibiotics have drastically decreased the mortality rate of many infectious agents. These remarkable successes in the control of infections came from knowing the causative agents of the diseases, followed by serendipitous discoveries of attenuated viruses and antibiotics. The discovery of DNA as genetic material and the understanding of how this information translates into specific phenotypes have changed the paradigm for developing new vaccines, drugs, and diagnostic tests. Knowledge of the mechanisms of immunity and mechanisms of action of drugs has led to new vaccines and new antimicrobial agents. The key to the acquisition of the knowledge of these mechanisms has been identifying the elemental causes (i.e., genes and their products) that mediate immunity and drug resistance. The identification of these genes is made possible by being able to transfer the genes or mutated forms of the genes into causative agents or surrogate hosts. Such an approach was limited in Mycobacterium tuberculosis by the difficulty of transferring genes or alleles into M. tuberculosis or a suitable surrogate mycobacterial host. The construction of shuttle phasmids—chimeric molecules that replicate in Escherichia coli as plasmids and in mycobacteria as mycobacteriophages—was instrumental in developing gene transfer systems for M. tuberculosis. This review will discuss M. tuberculosis genetic systems and their impact on tuberculosis research. “I had to know my enemy in order to prevail against him.”Nelson Mandela PMID:26105819

  16. ENERGY-TRANSFER SYSTEMS

    DOEpatents

    Thonemann, P.C.; Cowhig, W.T.; Davenport, P.A.

    1963-04-01

    This patent relates to the transfer of energy in a traveling electromagnetic wave to direct-current electrical energy in a gaseous medium. The traveling wave is generated by means of a radio-frequency oscillator connected across a capacitance-loaded helix wound around a sealed tube enclosing the gaseous medium. The traveling wave causes the electrons within the medium to drift towards one end of the tube. The direct current appearing across electrodes placed at each end of the tube is then used by some electrical means. (AEC)

  17. Power Transfer in Physical Systems.

    ERIC Educational Resources Information Center

    Kaeck, Jack A.

    1990-01-01

    Explores the power transfer using (1) a simple electric circuit consisting of a power source with internal resistance; (2) two different mechanical systems (gravity driven and constant force driven); (3) ecological examples; and (4) a linear motor. (YP)

  18. Horizontal Gene Transfer, Dispersal and Haloarchaeal Speciation

    PubMed Central

    Papke, R. Thane; Corral, Paulina; Ram-Mohan, Nikhil; de la Haba, Rafael R.; Sánchez-Porro, Cristina; Makkay, Andrea; Ventosa, Antonio

    2015-01-01

    The Halobacteria are a well-studied archaeal class and numerous investigations are showing how their diversity is distributed amongst genomes and geographic locations. Evidence indicates that recombination between species continuously facilitates the arrival of new genes, and within species, it is frequent enough to spread acquired genes amongst all individuals in the population. To create permanent independent diversity and generate new species, barriers to recombination are probably required. The data support an interpretation that rates of evolution (e.g., horizontal gene transfer and mutation) are faster at creating geographically localized variation than dispersal and invasion are at homogenizing genetic differences between locations. Therefore, we suggest that recurrent episodes of dispersal followed by variable periods of endemism break the homogenizing forces of intrapopulation recombination and that this process might be the principal stimulus leading to divergence and speciation in Halobacteria. PMID:25997110

  19. Plasmid and clonal interference during post horizontal gene transfer evolution.

    PubMed

    Bedhomme, S; Perez Pantoja, D; Bravo, I G

    2017-02-16

    Plasmids are nucleic acid molecules that can drive their own replication in a living cell. They can be transmitted horizontally and can thrive in the host cell to high-copy numbers. Plasmid replication and gene expression consume cellular resources and cells carrying plasmids incur fitness costs. But many plasmids carry genes that can be beneficial under certain conditions, allowing the cell to endure in the presence of antibiotics, toxins, competitors or parasites. Horizontal transfer of plasmid-encoded genes can thus instantaneously confer differential adaptation to local or transient selection conditions. This conflict between cellular fitness and plasmid spread sets the scene for multilevel selection processes. We have engineered a system to study the short-term evolutionary impact of different synonymous versions of a plasmid-encoded antibiotic resistance gene. Applying experimental evolution under different selection conditions and deep sequencing allowed us to show rapid local adaptation to the presence of antibiotic and to the specific version of the resistance gene transferred. We describe the presence of clonal interference at two different levels: at the within-cell level, because a single cell can carry several plasmids, and at the between-cell level, because a bacterial population may contain several clones carrying different plasmids and displaying different fitness in the presence/absence of antibiotic. Understanding the within-cell and between-cell dynamics of plasmids after horizontal gene transfer is essential to unravel the dense network of mobile elements underlying the worldwide threat to public health of antibiotic resistance.

  20. Horizontal gene transfer and the evolution of methanogenic pathways.

    PubMed

    Fournier, Greg

    2009-01-01

    Horizontal gene transfer (HGT) is a driving force in the evolution of metabolic pathways, allowing novel enzymatic functions that provide a selective advantage to be rapidly incorporated into an organism's physiology. Here, the role of two HGT events in the evolution of methanogenesis is described. First, the acetoclastic sub-pathway of methanogenesis is shown to have evolved via a transfer of the ackA and pta genes from a cellulolytic clostridia to a family of methanogenic archaea. Second, the system for encoding the amino acid pyrrolysine, used for the synthesis of enzymes for methanogenesis from methylamines, is shown to likely have evolved via transfer from an ancient, unknown, deeply branching organismal lineage.

  1. Therapeutic option of plasmid-DNA based gene transfer.

    PubMed

    Taniyama, Yoshiaki; Azuma, Junya; Kunugiza, Yasuo; Iekushi, Kazuma; Rakugi, Hiromi; Morishita, Ryuichi

    2012-01-01

    Gene therapy offers a novel approach for the prevention and treatment of a variety of diseases, but it is not yet a common method in clinical cases because of various problems. Viral vectors show high efficiency of gene transfer, but they have some problems with toxicity and immunity. On the other hand, plasmid deoxyribonucleic acid (DNA)-based gene transfer is very safe, but its efficiency is relatively low. Especially, plasmid DNA gene therapy is used for cardiovascular disease because plasmid DNA transfer is possible for cardiac or skeletal muscle. Clinical angiogenic gene therapy using plasmid DNA gene transfer has been attempted in patients with peripheral artery disease, but a phase III clinical trial did not show sufficient efficiency. In this situation, more efficient plasmid DNA gene transfer is needed all over the world. This review focuses on plasmid DNA gene transfer and its enhancement, including ultrasound with microbubbles, electroporation, hydrodynamic method, gene gun, jet injection, cationic lipids and cationic polymers.

  2. "Active" cancer immunotherapy by anti-Met antibody gene transfer.

    PubMed

    Vigna, Elisa; Pacchiana, Giovanni; Mazzone, Massimiliano; Chiriaco, Cristina; Fontani, Lara; Basilico, Cristina; Pennacchietti, Selma; Comoglio, Paolo M

    2008-11-15

    Gene therapy provides a still poorly explored opportunity to treat cancer by "active" immunotherapy as it enables the transfer of genes encoding antibodies directed against specific oncogenic proteins. By a bidirectional lentiviral vector, we transferred the cDNA encoding the heavy and light chains of a monoclonal anti-Met antibody (DN-30) to epithelial cancer cells. In vitro, the transduced cells synthesized and secreted correctly assembled antibodies with the expected high affinity, inducing down-regulation of the Met receptor and strong inhibition of the invasive growth response. The inhibitory activity resulted (a) from the interference of the antibody with the Met receptor intracellular processing ("cell autonomous activity," in cis) and (b) from the antibody-induced cleavage of Met expressed at the cell surface ("bystander effect," in trans). The monoclonal antibody gene transferred into live animals by systemic administration or by local intratumor delivery resulted in substantial inhibition of tumor growth. These data provide proof of concept both for targeting the Met receptor and for a gene transfer-based immunotherapy strategy.

  3. ASSEMBLY TRANSFER SYSTEM DESCRIPTION DOCUMENT

    SciTech Connect

    B. Gorpani

    2000-06-26

    The Assembly Transfer System (ATS) receives, cools, and opens rail and truck transportation casks from the Carrier/Cask Handling System (CCHS). The system unloads transportation casks consisting of bare Spent Nuclear Fuel (SNF) assemblies, single element canisters, and Dual Purpose Canisters (DPCs). For casks containing DPCs, the system opens the DPCs and unloads the SNF. The system stages the assemblies, transfer assemblies to and from fuel-blending inventory pools, loads them into Disposal Containers (DCs), temporarily seals and inerts the DC, decontaminates the DC and transfers it to the Disposal Container Handling System. The system also prepares empty casks and DPCs for off-site shipment. Two identical Assembly Transfer System lines are provided in the Waste Handling Building (WHB). Each line operates independently to handle the waste transfer throughput and to support maintenance operations. Each system line primarily consists of wet and dry handling areas. The wet handling area includes a cask transport system, cask and DPC preparation system, and a wet assembly handling system. The basket transport system forms the transition between the wet and dry handling areas. The dry handling area includes the dry assembly handling system, assembly drying system, DC preparation system, and DC transport system. Both the wet and dry handling areas are controlled by the control and tracking system. The system operating sequence begins with moving transportation casks to the cask preparation area. The cask preparation operations consist of cask cavity gas sampling, cask venting, cask cool-down, outer lid removal, and inner shield plug lifting fixture attachment. Casks containing bare SNF (no DPC) are filled with water and placed in the cask unloading pool. The inner shield plugs are removed underwater. For casks containing a DPC, the cask lid(s) is removed, and the DPC is penetrated, sampled, vented, and cooled. A DPC lifting fixture is attached and the cask is placed

  4. RANGE: Gene Transfer of Reversibly Controlled Polycistronic Genes

    PubMed Central

    Chen, Yiwei; Cao, Liji; Luo, Chonglin; Ditzel, Désirée AW; Peter, Jörg; Sprengel, Rolf

    2013-01-01

    We developed a single vector recombinant adeno-associated viral (rAAV) expression system for spatial and reversible control of polycistronic gene expression. Our approach (i) integrates the advantages of the tetracycline (Tet)-controlled transcriptional silencer tTSKid and the self-cleaving 2A peptide bridge, (ii) combines essential regulatory components as an autoregulatory loop, (iii) simplifies the gene delivery scheme, and (iv) regulates multiple genes in a synchronized manner. Controlled by an upstream Tet-responsive element (TRE), both the ubiquitous chicken β-actin promoter (CAG) and the neuron-specific synapsin-1 promoter (Syn) could regulate expression of tTSKid together with two 2A-linked reporter genes. Transduction in vitro exhibited maximally 50-fold regulation by doxycycline (Dox). Determined by gene delivery method as well as promoter, highly specific tissues were transduced in vivo. Bioluminescence imaging (BLI) visualized reversible “ON/OFF” gene switches over repeated “Doxy-Cycling” in living mice. Thus, the reversible rAAV-mediated N-cistronic gene expression system, termed RANGE, may serve as a versatile tool to achieve reversible polycistronic gene regulation for the study of gene function as well as gene therapy. PMID:23571608

  5. Promoters and serotypes: targeting of adeno-associated virus vectors for gene transfer in the rat central nervous system in vitro and in vivo.

    PubMed

    Shevtsova, Z; Malik, J M I; Michel, U; Bähr, M; Kügler, S

    2005-01-01

    The brain parenchyma consists of several different cell types, such as neurones, astrocytes, microglia, oligodendroglia and epithelial cells, which are morphologically and functionally intermingled in highly complex three-dimensional structures. These different cell types are also present in cultures of brain cells prepared to serve as model systems of CNS physiology. Gene transfer, either in a therapeutic attempt or in basic research, is a fascinating and promising tool to manipulate both the complex physiology of the brain and that of isolated neuronal cells. Viral vectors based on the parvovirus, adeno-associated virus (AAV), have emerged as powerful transgene delivery vehicles. Here we describe highly efficient targeting of AAV vectors to either neurones or astrocytes in cultured primary brain cell cultures. We also show that transcriptional targeting can be achieved by the use of small promoters, significantly boosting the transgene capacity of the recombinant viral genome. However, we also demonstrate that successful targeting of a vector in vitro does not necessarily imply that the same targeting works in the adult brain. Cross-packaging the AAV-2 genome in capsids of other serotypes adds additional benefits to this vector system. In the brain, the serotype-5 capsid allows for drastically increased spread of the recombinant vector as compared to the serotype-2 capsid. Finally, we emphasize the optimal targeting approach, in which the natural tropism of a vector for a specific cell type is employed. Taken together, these data demonstrate the flexibility which AAV-based vector systems offer in physiological research.

  6. Reducible cationic lipids for gene transfer.

    PubMed Central

    Wetzer, B; Byk, G; Frederic, M; Airiau, M; Blanche, F; Pitard, B; Scherman, D

    2001-01-01

    One of the main challenges of gene therapy remains the increase of gene delivery into eukaryotic cells. We tested whether intracellular DNA release, an essential step for gene transfer, could be facilitated by using reducible cationic DNA-delivery vectors. For this purpose, plasmid DNA was complexed with cationic lipids bearing a disulphide bond. This reduction-sensitive linker is expected to be reduced and cleaved in the reducing milieu of the cytoplasm, thus potentially improving DNA release and consequently transfection. The DNA--disulphide-lipid complexation was monitored by ethidium bromide exclusion, and the size of complexes was determined by dynamic light scattering. It was found that the reduction kinetics of disulphide groups in DNA--lipid complexes depended on the position of the disulphide linker within the lipid molecule. Furthermore, the internal structure of DNA--lipid particles was examined by small-angle X-ray scattering before and after lipid reduction. DNA release from lipid complexes was observed after the reduction of disulphide bonds of several lipids. Cell-transfection experiments suggested that complexes formed with selected reducible lipids resulted in up to 1000-fold higher reporter-gene activity, when compared with their analogues without disulphide bonds. In conclusion, reduction-sensitive groups introduced into cationic lipid backbones potentially allow enhanced DNA release from DNA--lipid complexes after intracellular reduction and represent a tool for improved vectorization. PMID:11389682

  7. Reducible cationic lipids for gene transfer.

    PubMed

    Wetzer, B; Byk, G; Frederic, M; Airiau, M; Blanche, F; Pitard, B; Scherman, D

    2001-06-15

    One of the main challenges of gene therapy remains the increase of gene delivery into eukaryotic cells. We tested whether intracellular DNA release, an essential step for gene transfer, could be facilitated by using reducible cationic DNA-delivery vectors. For this purpose, plasmid DNA was complexed with cationic lipids bearing a disulphide bond. This reduction-sensitive linker is expected to be reduced and cleaved in the reducing milieu of the cytoplasm, thus potentially improving DNA release and consequently transfection. The DNA--disulphide-lipid complexation was monitored by ethidium bromide exclusion, and the size of complexes was determined by dynamic light scattering. It was found that the reduction kinetics of disulphide groups in DNA--lipid complexes depended on the position of the disulphide linker within the lipid molecule. Furthermore, the internal structure of DNA--lipid particles was examined by small-angle X-ray scattering before and after lipid reduction. DNA release from lipid complexes was observed after the reduction of disulphide bonds of several lipids. Cell-transfection experiments suggested that complexes formed with selected reducible lipids resulted in up to 1000-fold higher reporter-gene activity, when compared with their analogues without disulphide bonds. In conclusion, reduction-sensitive groups introduced into cationic lipid backbones potentially allow enhanced DNA release from DNA--lipid complexes after intracellular reduction and represent a tool for improved vectorization.

  8. Horizontal gene transfer from Agrobacterium to plants

    PubMed Central

    Matveeva, Tatiana V.; Lutova, Ludmila A.

    2014-01-01

    Most genetic engineering of plants uses Agrobacterium mediated transformation to introduce novel gene content. In nature, insertion of T-DNA in the plant genome and its subsequent transfer via sexual reproduction has been shown in several species in the genera Nicotiana and Linaria. In these natural examples of horizontal gene transfer from Agrobacterium to plants, the T-DNA donor is assumed to be a mikimopine strain of A. rhizogenes. A sequence homologous to the T-DNA of the Ri plasmid of Agrobacterium rhizogenes was found in the genome of untransformed Nicotiana glauca about 30 years ago, and was named “cellular T-DNA” (cT-DNA). It represents an imperfect inverted repeat and contains homologs of several T-DNA oncogenes (NgrolB, NgrolC, NgORF13, NgORF14) and an opine synthesis gene (Ngmis). A similar cT-DNA has also been found in other species of the genus Nicotiana. These presumably ancient homologs of T-DNA genes are still expressed, indicating that they may play a role in the evolution of these plants. Recently T-DNA has been detected and characterized in Linaria vulgaris and L. dalmatica. In Linaria vulgaris the cT-DNA is present in two copies and organized as a tandem imperfect direct repeat, containing LvORF2, LvORF3, LvORF8, LvrolA, LvrolB, LvrolC, LvORF13, LvORF14, and the Lvmis genes. All L. vulgaris and L. dalmatica plants screened contained the same T-DNA oncogenes and the mis gene. Evidence suggests that there were several independent T-DNA integration events into the genomes of these plant genera. We speculate that ancient plants transformed by A. rhizogenes might have acquired a selective advantage in competition with the parental species. Thus, the events of T-DNA insertion in the plant genome might have affected their evolution, resulting in the creation of new plant species. In this review we focus on the structure and functions of cT-DNA in Linaria and Nicotiana and discuss their possible evolutionary role. PMID:25157257

  9. Lateral gene transfer, bacterial genome evolution, and the Anthropocene.

    PubMed

    Gillings, Michael R

    2017-02-01

    Lateral gene transfer (LGT) has significantly influenced bacterial evolution since the origins of life. It helped bacteria generate flexible, mosaic genomes and enables individual cells to rapidly acquire adaptive phenotypes. In turn, this allowed bacteria to mount strong defenses against human attempts to control their growth. The widespread dissemination of genes conferring resistance to antimicrobial agents has precipitated a crisis for modern medicine. Our actions can promote increased rates of LGT and also provide selective forces to fix such events in bacterial populations. For instance, the use of selective agents induces the bacterial SOS response, which stimulates LGT. We create hotspots for lateral transfer, such as wastewater systems, hospitals, and animal production facilities. Conduits of gene transfer between humans and animals ensure rapid dissemination of recent transfer events, as does modern transport and globalization. As resistance to antibacterial compounds becomes universal, there is likely to be increasing selection pressure for phenotypes with adverse consequences for human welfare, such as enhanced virulence, pathogenicity, and transmission. Improved understanding of the ecology of LGT could help us devise strategies to control this fundamental evolutionary process.

  10. Kidney-specific transposon-mediated gene transfer in vivo.

    PubMed

    Woodard, Lauren E; Cheng, Jizhong; Welch, Richard C; Williams, Felisha M; Luo, Wentian; Gewin, Leslie S; Wilson, Matthew H

    2017-03-20

    Methods enabling kidney-specific gene transfer in adult mice are needed to develop new therapies for kidney disease. We attempted kidney-specific gene transfer following hydrodynamic tail vein injection using the kidney-specific podocin and gamma-glutamyl transferase promoters, but found expression primarily in the liver. In order to achieve kidney-specific transgene expression, we tested direct hydrodynamic injection of a DNA solution into the renal pelvis and found that luciferase expression was strong in the kidney and absent from extra-renal tissues. We observed heterogeneous, low-level transfection of the collecting duct, proximal tubule, distal tubule, interstitial cells, and rarely glomerular cells following injection. To assess renal injury, we performed the renal pelvis injections on uninephrectomised mice and found that their blood urea nitrogen was elevated at two days post-transfer but resolved within two weeks. Although luciferase expression quickly decreased following renal pelvis injection, the use of the piggyBac transposon system improved long-term expression. Immunosuppression with cyclophosphamide stabilised luciferase expression, suggesting immune clearance of the transfected cells occurs in immunocompetent animals. Injection of a transposon expressing erythropoietin raised the haematocrit, indicating that the developed injection technique can elicit a biologic effect in vivo. Hydrodynamic renal pelvis injection enables transposon mediated-kidney specific gene transfer in adult mice.

  11. Kidney-specific transposon-mediated gene transfer in vivo

    PubMed Central

    Woodard, Lauren E.; Cheng, Jizhong; Welch, Richard C.; Williams, Felisha M.; Luo, Wentian; Gewin, Leslie S.; Wilson, Matthew H.

    2017-01-01

    Methods enabling kidney-specific gene transfer in adult mice are needed to develop new therapies for kidney disease. We attempted kidney-specific gene transfer following hydrodynamic tail vein injection using the kidney-specific podocin and gamma-glutamyl transferase promoters, but found expression primarily in the liver. In order to achieve kidney-specific transgene expression, we tested direct hydrodynamic injection of a DNA solution into the renal pelvis and found that luciferase expression was strong in the kidney and absent from extra-renal tissues. We observed heterogeneous, low-level transfection of the collecting duct, proximal tubule, distal tubule, interstitial cells, and rarely glomerular cells following injection. To assess renal injury, we performed the renal pelvis injections on uninephrectomised mice and found that their blood urea nitrogen was elevated at two days post-transfer but resolved within two weeks. Although luciferase expression quickly decreased following renal pelvis injection, the use of the piggyBac transposon system improved long-term expression. Immunosuppression with cyclophosphamide stabilised luciferase expression, suggesting immune clearance of the transfected cells occurs in immunocompetent animals. Injection of a transposon expressing erythropoietin raised the haematocrit, indicating that the developed injection technique can elicit a biologic effect in vivo. Hydrodynamic renal pelvis injection enables transposon mediated-kidney specific gene transfer in adult mice. PMID:28317878

  12. Gene duplication and transfer events in plant mitochondria genome

    SciTech Connect

    Xiong Aisheng Peng Rihe; Zhuang Jing; Gao Feng; Zhu Bo; Fu Xiaoyan; Xue Yong; Jin Xiaofen; Tian Yongsheng; Zhao Wei; Yao Quanhong

    2008-11-07

    Gene or genome duplication events increase the amount of genetic material available to increase the genomic, and thereby phenotypic, complexity of organisms during evolution. Gene duplication and transfer events have been important to molecular evolution in all three domains of life, and may be the first step in the emergence of new gene functions. Gene transfer events have been proposed as another accelerator of evolution. The duplicated gene or genome, mainly nuclear, has been the subject of several recent reviews. In addition to the nuclear genome, organisms have organelle genomes, including mitochondrial genome. In this review, we briefly summarize gene duplication and transfer events in the plant mitochondrial genome.

  13. Horizontal gene transfer: A critical view

    PubMed Central

    Kurland, C. G.; Canback, B.; Berg, Otto G.

    2003-01-01

    It has been suggested that horizontal gene transfer (HGT) is the “essence of phylogeny.” In contrast, much data suggest that this is an exaggeration resulting in part from a reliance on inadequate methods to identify HGT events. In addition, the assumption that HGT is a ubiquitous influence throughout evolution is questionable. Instead, rampant global HGT is likely to have been relevant only to primitive genomes. In modern organisms we suggest that both the range and frequencies of HGT are constrained most often by selective barriers. As a consequence those HGT events that do occur most often have little influence on genome phylogeny. Although HGT does occur with important evolutionary consequences, classical Darwinian lineages seem to be the dominant mode of evolution for modern organisms. PMID:12902542

  14. uv excision-repair gene transfer in Chinese hamster ovary (CHO) cells

    SciTech Connect

    MacInnes, M.A.; Bingham, J.M.; Strniste, G.F.; Thompson, L.H.

    1983-01-01

    uvc-sensitive mutants of CHO cells provide a model system for molecular studies of DNA repair. We present our recent results which show that these mutants are competent recipients for plasmid marker gene transfer and incorporation of a putative CHO repair gene. The applicability and advantages of this system for interspecies human repair gene identification are discussed.

  15. Horizontal transfer of expressed genes in a parasitic flowering plant

    PubMed Central

    2012-01-01

    Background Recent studies have shown that plant genomes have potentially undergone rampant horizontal gene transfer (HGT). In plant parasitic systems HGT appears to be facilitated by the intimate physical association between the parasite and its host. HGT in these systems has been invoked when a DNA sequence obtained from a parasite is placed phylogenetically very near to its host rather than with its closest relatives. Studies of HGT in parasitic plants have relied largely on the fortuitous discovery of gene phylogenies that indicate HGT, and no broad systematic search for HGT has been undertaken in parasitic systems where it is most expected to occur. Results We analyzed the transcriptomes of the holoparasite Rafflesia cantleyi Solms-Laubach and its obligate host Tetrastigma rafflesiae Miq. using phylogenomic approaches. Our analyses show that several dozen actively transcribed genes, most of which appear to be encoded in the nuclear genome, are likely of host origin. We also find that hundreds of vertically inherited genes (VGT) in this parasitic plant exhibit codon usage properties that are more similar to its host than to its closest relatives. Conclusions Our results establish for the first time a substantive number of HGTs in a plant host-parasite system. The elevated rate of unidirectional host-to- parasite gene transfer raises the possibility that HGTs may provide a fitness benefit to Rafflesia for maintaining these genes. Finally, a similar convergence in codon usage of VGTs has been shown in microbes with high HGT rates, which may help to explain the increase of HGTs in these parasitic plants. PMID:22681756

  16. Optical gene transfer by femtosecond laser pulses

    NASA Astrophysics Data System (ADS)

    Konig, Karsten; Riemann, Iris; Tirlapur, Uday K.

    2003-07-01

    Targeted transfection of cells is an important technique for gene therapy and related biomedical applications. We delineate how high-intensity (1012 W/cm2) near-infrared (NIR) 80 MHz nanojoule femtosecond laser pulses can create highly localised membrane perforations within a minute focal volume, enabling non-invasive direct transfection of mammalian cells with DNA. We suspended Chinese hamster ovarian (CHO), rat kangaroo kidney epithelial (PtK2) and rat fibroblast cells in 0.5 ml culture medium in a sterile miniaturized cell chamber (JenLab GmbH, Jena, Germany) containing 0.2 μg plasmid DNA vector pEGFP-N1 (4.7 kb), which codes for green fluorescent protein (GFP). The NIR laser beam was introduced into a femtosecond laser scanning microscope (JenLab GmbH, Jena, Germany; focussed on the edge of the cell membrane of a target cell for 16 ms. The integration and expression efficiency of EGFP were assessed in situ by two-photon fluorescence-lifetime imaging using time-correlated single photon counting. The unique capability to transfer foreign DNA safely and efficiently into specific cell types (including stem cells), circumventing mechanical, electrical or chemical means, will have many applications, such as targeted gene therapy and DNA vaccination.

  17. Plant transformation via pollen tube-mediated gene transfer

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Genetic transformation using foreign genes and the subsequent development of transgenic plants has been employed to develop enhanced elite germplasm. Although some skepticism exits regarding pollen tube-mediated gene transfer (PTT), reports demonstrating improved transformation efficiency with PTT ...

  18. Gene Transfer & Hybridization Studies in Hyperthermophilic Species

    SciTech Connect

    Nelson, Karen E.

    2005-10-14

    A. ABSTRACT The importance of lateral gene transfer (LGT) in the evolution of microbial species has become increasingly evident with each completed microbial genome sequence. Most significantly, the genome of Thermotoga maritima MSB8, a hyperthermophilic bacterium isolated by Karl Stetter and workers from Vulcano Italy in 1986, and sequenced at The Institute for Genomic Research (TIGR) in Rockville Maryland in 1999, revealed extensive LGT between % . this bacterium and members of the archaeal domain (in particular Archaeoglobus fulgidus, and Pyracoccus frcriosus species). Based on whole genome comparisons, it was estimated that 24% of the genetic information in this organism was acquired by genetic exchange with archaeal species, Independent analyses including periodicity analysis of the T. maritimu genomic DNA sequence, phylogenetic reconstruction based on genes that appear archaeal-like, and codon and amino acid usage, have provided additional evidence for LGT between T. maritima and the archaea. More recently, DiRuggiero and workers have identified a very recent LGT event between two genera of hyperthermophilic archaea, where a nearly identical DNA fragment of 16 kb in length flanked by insertion sequence (IS) elements, exists. Undoubtedly, additional examples of LGT will be identified as more microbial genomes are completed. For the present moment however, the genome sequence of T. maritima and other hyperthermophiles including P. furiosus, Pyrococcus horikoshii, Pyrococcus abyssi, A. fulgidus, and Aquifex aeolicus, have significantly increased out awareness of evolution being a web of life rather than a tree of life, as suggested by single gene phylogenies. In this proposal, we will aim to determine the extent of LGT across the hyperthemophiles, employing iY maritima as the model organism. A variety of biochemical techniques and phylogenetic reconstructions will allow for a detailed and thorough characterization of the extent of LGT in this species. The

  19. Plasmid DNA-based gene transfer with ultrasound and microbubbles.

    PubMed

    Taniyama, Yoshiaki; Azuma, Junya; Rakugi, Hiromi; Morishita, Ryuichi

    2011-12-01

    Gene therapy offers a novel approach for the prevention and treatment of a variety of diseases, but it is not yet a common option in the real world because of various problems. Viral vectors show high efficiency of gene transfer, but they have some problems with toxicity and immunity. On the other hand, plasmid DNA-based gene transfer is very safe, but its efficiency is relatively low. Especially, plasmid DNA gene therapy is used for cardiovascular disease because plasmid DNA transfer is possible for cardiac or skeletal muscle. Clinical angiogenic gene therapy using plasmid DNA gene transfer has been attempted in patients with peripheral artery disease, but a Phase III clinical trial did not show sufficient efficiency. Recently, a Phase III clinical trial of hepatocyte growth factor gene therapy in peripheral artery disease (PAD) showed improvement of ischemic ulcers, but it could not salvage limbs from amputation. In addition, a Phase I/II clinical study of fibroblast growth factor gene therapy in PAD extended amputation-free survival, but it seemed to fail in Phase III. In this situation, we and others have developed plasmid DNA-based gene transfer using ultrasound with microbubbles to enhance its efficiency while maintaining safety. Ultrasound-mediated gene transfer has been reported to augment the gene transfer efficiency and select the target organ using cationic microbubble phospholipids which bind negatively charged DNA. Ultrasound with microbubblesis likely to create new therapeutic options inavariety of diseases.

  20. Dynamic monitoring of horizontal gene transfer in soil

    NASA Astrophysics Data System (ADS)

    Cheng, H. Y.; Masiello, C. A.; Silberg, J. J.; Bennett, G. N.

    2015-12-01

    Soil microbial gene expression underlies microbial behaviors (phenotypes) central to many aspects of C, N, and H2O cycling. However, continuous monitoring of microbial gene expression in soils is challenging because genetically-encoded reporter proteins widely used in the lab are difficult to deploy in soil matrices: for example, green fluorescent protein cannot be easily visualized in soils, even in the lab. To address this problem we have developed a reporter protein that releases small volatile gases. Here, we applied this gas reporter in a proof-of-concept soil experiment, monitoring horizontal gene transfer, a microbial activity that alters microbial genotypes and phenotypes. Horizontal gene transfer is central to bacterial evolution and adaptation and is relevant to problems such as the spread of antibiotic resistance, increasing metal tolerance in superfund sites, and bioremediation capability of bacterial consortia. This process is likely to be impacted by a number of matrix properties not well-represented in the petri dish, such as microscale variations in water, nutrients, and O2, making petri-dish experiments a poor proxy for environmental processes. We built a conjugation system using synthetic biology to demonstrate the use of gas-reporting biosensors in safe, lab-based biogeochemistry experiments, and here we report the use of these sensors to monitor horizontal gene transfer in soils. Our system is based on the F-plasmid conjugation in Escherichia coli. We have found that the gas signal reports on the number of cells that acquire F-plasmids (transconjugants) in a loamy Alfisol collected from Kellogg Biological Station. We will report how a gas signal generated by transconjugants varies with the number of F-plasmid donor and acceptor cells seeded in a soil, soil moisture, and soil O2 levels.

  1. Evaluation of biolistic gene transfer methods in vivo using non-invasive bioluminescent imaging techniques

    PubMed Central

    2011-01-01

    Background Gene therapy continues to hold great potential for treating many different types of disease and dysfunction. Safe and efficient techniques for gene transfer and expression in vivo are needed to enable gene therapeutic strategies to be effective in patients. Currently, the most commonly used methods employ replication-defective viral vectors for gene transfer, while physical gene transfer methods such as biolistic-mediated ("gene-gun") delivery to target tissues have not been as extensively explored. In the present study, we evaluated the efficacy of biolistic gene transfer techniques in vivo using non-invasive bioluminescent imaging (BLI) methods. Results Plasmid DNA carrying the firefly luciferase (LUC) reporter gene under the control of the human Cytomegalovirus (CMV) promoter/enhancer was transfected into mouse skin and liver using biolistic methods. The plasmids were coupled to gold microspheres (1 μm diameter) using different DNA Loading Ratios (DLRs), and "shot" into target tissues using a helium-driven gene gun. The optimal DLR was found to be in the range of 4-10. Bioluminescence was measured using an In Vivo Imaging System (IVIS-50) at various time-points following transfer. Biolistic gene transfer to mouse skin produced peak reporter gene expression one day after transfer. Expression remained detectable through four days, but declined to undetectable levels by six days following gene transfer. Maximum depth of tissue penetration following biolistic transfer to abdominal skin was 200-300 μm. Similarly, biolistic gene transfer to mouse liver in vivo also produced peak early expression followed by a decline over time. In contrast to skin, however, liver expression of the reporter gene was relatively stable 4-8 days post-biolistic gene transfer, and remained detectable for nearly two weeks. Conclusions The use of bioluminescence imaging techniques enabled efficient evaluation of reporter gene expression in vivo. Our results demonstrate that

  2. AAV-Mediated Gene Transfer to Dorsal Root Ganglion.

    PubMed

    Yu, Hongwei; Fischer, Gregory; Hogan, Quinn H

    2016-01-01

    Transferring genetic molecules into the peripheral sensory nervous system to manipulate nociceptive pathophysiology is a powerful approach for experimental modulation of sensory signaling and potentially for translation into therapy for chronic pain. This can be efficiently achieved by the use of recombinant adeno-associated virus (rAAV) in conjunction with nociceptor-specific regulatory transgene cassettes. Among different routes of delivery, direct injection into the dorsal root ganglia (DRGs) offers the most efficient AAV-mediated gene transfer selectively into the peripheral sensory nervous system. Here, we briefly discuss the advantages and applications of intraganglionic microinjection, and then provide a detailed approach for DRG injection, including a list of the necessary materials and description of a method for performing DRG microinjection experiments. We also discuss our experience with several adeno-associated virus (AAV) options for in vivo transgene expression in DRG neurons.

  3. In vivo Cytokine Gene Transfer by Gene Gun Reduces Tumor Growth in Mice

    NASA Astrophysics Data System (ADS)

    Sun, Wenn H.; Burkholder, Joseph K.; Sun, Jian; Culp, Jerilyn; Turner, Joel; Lu, Xing G.; Pugh, Thomas D.; Ershler, William B.; Yang, Ning-Sun

    1995-03-01

    Implantation of tumor cells modified by in vitro cytokine gene transfer has been shown by many investigators to result in potent in vivo antitumor activities in mice. Here we describe an approach to tumor immunotherapy utilizing direct transfection of cytokine genes into tumorbearing animals by particle-mediated gene transfer. In vivo transfection of the human interleukin 6 gene into the tumor site reduced methylcholanthrene-induced fibrosarcoma growth, and a combination of murine tumor necrosis factor α and interferon γ genes inhibited growth of a renal carcinoma tumor model (Renca). In addition, treatment with murine interleukin 2 and interferon γ genes prolonged the survival of Renca tumor-bearing mice and resulted in tumor eradication in 25% of the test animals. Transgene expression was demonstrated in treated tissues by ELISA and immunohistochemical analysis. Significant serum levels of interleukin 6 and interferon γ were detected, demonstrating effective secretion of transgenic proteins from treated skin into the bloodstream. This in vivo cytokine gene therapy approach provides a system for evaluating the antitumor properties of various cytokines in different tumor models and has potential utility for human cancer gene therapy.

  4. Problems associated with gene transfer and opportunities for microgravity environments

    SciTech Connect

    Tennessen, D.J.

    1997-01-01

    The method of crop improvement by gene transfer is becoming increasingly routine with transgenic foods and ornamental crops now being marketed to consumers. However, biological processes of plants, and the physical barriers of current protocols continue to limit the application of gene transfer in many commercial crops. The goal of this paper is to outline the current limitations of gene transfer and to hypothesize possible opportunities for use of microgravity to overcome such limitations. The limitations detailed in this paper include host-range specificity of {ital Agrobacterium} mediated transformation, probability of gene insertion, position effects of the inserted genes, gene copy number, stability of foreign gene expression in host plants, and regeneration of recalcitrant plant species. Microgravity offers an opportunity for gene transfer where cell growth kinetics, DNA synthesis, and genetic recombination rates can be altered. Such biological conditions may enhance the ability for recombination of reporter genes and other genes of interest to agriculture. Proposed studies would be useful for understanding instability of foreign gene expression and may lead to stable transformed plants. Other aspects of gene transfer in microgravity are discussed. {copyright} {ital 1997 American Institute of Physics.}

  5. Problems associated with gene transfer and opportunities for microgravity environments

    NASA Astrophysics Data System (ADS)

    Tennessen, Daniel J.

    1997-01-01

    The method of crop improvement by gene transfer is becoming increasingly routine with transgenic foods and ornamental crops now being marketed to consumers. However, biological processes of plants, and the physical barriers of current protocols continue to limit the application of gene transfer in many commercial crops. The goal of this paper is to outline the current limitations of gene transfer and to hypothesize possible opportunities for use of microgravity to overcome such limitations. The limitations detailed in this paper include host-range specificity of Agrobacterium mediated transformation, probability of gene insertion, position effects of the inserted genes, gene copy number, stability of foreign gene expression in host plants, and regeneration of recalcitrant plant species. Microgravity offers an opportunity for gene transfer where cell growth kinetics, DNA synthesis, and genetic recombination rates can be altered. Such biological conditions may enhance the ability for recombination of reporter genes and other genes of interest to agriculture. Proposed studies would be useful for understanding instability of foreign gene expression and may lead to stable transformed plants. Other aspects of gene transfer in microgravity are discussed.

  6. Optical fiber data transfer system

    NASA Technical Reports Server (NTRS)

    Mcmillan, S. H.

    1988-01-01

    This Phase 2 effort applies the results of Phase 1 to design and fabricate an optical slip ring system for a helicopter rotor blade/wind tunnel application. In this application, there are two assemblies: one on the rotating portion of the mechanical system, one on the stationary portion. The assembly on the rotating portion digitizes and encodes 128 transducer signals from various parts of the blade, and optically transfers data across the noncontacting coupling. Two complete identical independent channels are provided. On the stationary side, the signals are decoded and one channel is transmitted in digital form to a computer for recording and analysis. The second channel reconstructs the analog transducer signals for real time observation. In the opposite direction, eight signal channels enable control signals to be passed from the stationary to the rotating part of the system. Power to the rotor mounted electronics is supplied via power slip rings. The advantages of the optical over the traditional electro-mechanical slip ring method of data transfer across a rotating joint are long life, low-maintenance, immunity to crosstalk, and wider bandwidth. Successful completion of this effort demonstrated that this method is practical and reliable, and can be implemented under difficult conditions of available space, power, environment, and stringent performance and equipment life requirements.

  7. Therapeutic antibody gene transfer: an active approach to passive immunity.

    PubMed

    Bakker, Joost M; Bleeker, Wim K; Parren, Paul W H I

    2004-09-01

    Advances in gene transfer approaches are enabling the possibility of applying therapeutic antibodies using DNA. In particular gene transfer in combination with electroporation is promising and can result in generating in vivo antibody concentrations in the low therapeutic range. However, several important problems need to be dealt with before antibody gene transfer can become a valuable supplement to the current therapies. As antibody production following gene transfer is difficult to control, the danger of inducing autoimmune conditions or uncontrollable side effects occurs in cases in which autologous antigens are targeted. It is suggested that the most promising area of application therefore appears to be infectious disease in which heterologous antigens are targeted and concerns for long-term antibody exposure are minimal. Finally, genes encoding fully human antibodies will enhance long-term expression and decrease problems linked to immunogenicity.

  8. Waste Feed Delivery Transfer System Analysis

    SciTech Connect

    JULYK, L.J.

    2000-05-05

    This document provides a documented basis for the required design pressure rating and pump pressure capacity of the Hanford Site waste-transfer system in support of the waste feed delivery to the privatization contractor for vitrification. The scope of the analysis includes the 200 East Area double-shell tank waste transfer pipeline system and the associated transfer system pumps for a11 Phase 1B and Phase 2 waste transfers from AN, AP, AW, AY, and A2 Tank Farms.

  9. The power of phylogenetic approaches to detect horizontally transferred genes

    PubMed Central

    Poptsova, Maria S; Gogarten, J Peter

    2007-01-01

    Background Horizontal gene transfer plays an important role in evolution because it sometimes allows recipient lineages to adapt to new ecological niches. High genes transfer frequencies were inferred for prokaryotic and early eukaryotic evolution. Does horizontal gene transfer also impact phylogenetic reconstruction of the evolutionary history of genomes and organisms? The answer to this question depends at least in part on the actual gene transfer frequencies and on the ability to weed out transferred genes from further analyses. Are the detected transfers mainly false positives, or are they the tip of an iceberg of many transfer events most of which go undetected by current methods? Results Phylogenetic detection methods appear to be the method of choice to infer gene transfers, especially for ancient transfers and those followed by orthologous replacement. Here we explore how well some of these methods perform using in silico transfers between the terminal branches of a gamma proteobacterial, genome based phylogeny. For the experiments performed here on average the AU test at a 5% significance level detects 90.3% of the transfers and 91% of the exchanges as significant. Using the Robinson-Foulds distance only 57.7% of the exchanges and 60% of the donations were identified as significant. Analyses using bipartition spectra appeared most successful in our test case. The power of detection was on average 97% using a 70% cut-off and 94.2% with 90% cut-off for identifying conflicting bipartitions, while the rate of false positives was below 4.2% and 2.1% for the two cut-offs, respectively. For all methods the detection rates improved when more intervening branches separated donor and recipient. Conclusion Rates of detected transfers should not be mistaken for the actual transfer rates; most analyses of gene transfers remain anecdotal. The method and significance level to identify potential gene transfer events represent a trade-off between the frequency of erroneous

  10. Smart crane ammunition transfer system

    SciTech Connect

    Bradley, E.C.; Killough, S.M.; Rowe, J.C.

    1995-03-01

    The purpose of the Smart Crane Ammunition Transfer System (SCATS) project is to demonstrate robotic/telerobotic controls technology for a mobile articulated crane for missile/ munitions handling, delivery, and reload. Missile resupply and reload have been manually intensive operations up to this time. Currently, reload missiles are delivered by truck to the site of the launcher. A crew of four to five personnel reloads the missiles from the truck to the launcher using a hydraulic-powered crane. The missiles are handled carefully for the safety of the missiles and personnel. Numerous steps are required in the reload process and the entire reload operation can take over 1 h for some missile systems. Recent U.S. Army directives require the entire operation to be accomplished in a fraction of that time. Current requirements for the development of SCATS are being based primarily on reloading Patriot missiles. The planned development approach will integrate robotic control and sensor technology with a commercially available hydraulic articulated crane. SCATS is being developed with commercially available hardware as much as possible. Development plans include adding a 3-D.F. end effector with a grapple to the articulating crane; closed-loop position control for the crane and end effector; digital microprocessor control of crane functions; simplified operator interface; and operating modes which include rectilinear movement, obstacle avoidance, and partial automated operation. The planned development will include progressive technology demonstrations. Ultimate plans are for this technology to be transferred and utilized in the military fielding process.

  11. A recently transferred cluster of bacterial genes in Trichomonas vaginalis - lateral gene transfer and the fate of acquired genes

    PubMed Central

    2014-01-01

    Background Lateral Gene Transfer (LGT) has recently gained recognition as an important contributor to some eukaryote proteomes, but the mechanisms of acquisition and fixation in eukaryotic genomes are still uncertain. A previously defined norm for LGTs in microbial eukaryotes states that the majority are genes involved in metabolism, the LGTs are typically localized one by one, surrounded by vertically inherited genes on the chromosome, and phylogenetics shows that a broad collection of bacterial lineages have contributed to the transferome. Results A unique 34 kbp long fragment with 27 clustered genes (TvLF) of prokaryote origin was identified in the sequenced genome of the protozoan parasite Trichomonas vaginalis. Using a PCR based approach we confirmed the presence of the orthologous fragment in four additional T. vaginalis strains. Detailed sequence analyses unambiguously suggest that TvLF is the result of one single, recent LGT event. The proposed donor is a close relative to the firmicute bacterium Peptoniphilus harei. High nucleotide sequence similarity between T. vaginalis strains, as well as to P. harei, and the absence of homologs in other Trichomonas species, suggests that the transfer event took place after the radiation of the genus Trichomonas. Some genes have undergone pseudogenization and degradation, indicating that they may not be retained in the future. Functional annotations reveal that genes involved in informational processes are particularly prone to degradation. Conclusions We conclude that, although the majority of eukaryote LGTs are single gene occurrences, they may be acquired in clusters of several genes that are subsequently cleansed of evolutionarily less advantageous genes. PMID:24898731

  12. LATERAL GENE TRANSFER AND THE HISTORY OF BACTERIAL GENOMES

    SciTech Connect

    Howard Ochman

    2006-02-22

    The aims of this research were to elucidate the role and extent of lateral transfer in the differentiation of bacterial strains and species, and to assess the impact of gene transfer on the evolution of bacterial genomes. The ultimate goal of the project is to examine the dynamics of a core set of protein-coding genes (i.e., those that are distributed universally among Bacteria) by developing conserved primers that would allow their amplification and sequencing in any bacterial taxa. In addition, we adopted a bioinformatic approach to elucidate the extent of lateral gene transfer in sequenced genome.

  13. Horizontal gene transfer of an entire metabolic pathway between a eukaryotic alga and its DNA virus

    PubMed Central

    Monier, Adam; Pagarete, António; de Vargas, Colomban; Allen, Michael J.; Read, Betsy; Claverie, Jean-Michel; Ogata, Hiroyuki

    2009-01-01

    Interactions between viruses and phytoplankton, the main primary producers in the oceans, affect global biogeochemical cycles and climate. Recent studies are increasingly revealing possible cases of gene transfers between cyanobacteria and phages, which might have played significant roles in the evolution of cyanobacteria/phage systems. However, little has been documented about the occurrence of horizontal gene transfer in eukaryotic phytoplankton/virus systems. Here we report phylogenetic evidence for the transfer of seven genes involved in the sphingolipid biosynthesis pathway between the cosmopolitan eukaryotic microalga Emiliania huxleyi and its large DNA virus EhV. PCR assays indicate that these genes are prevalent in E. huxleyi and EhV strains isolated from different geographic locations. Patterns of protein and gene sequence conservation support that these genes are functional in both E. huxleyi and EhV. This is the first clear case of horizontal gene transfer of multiple functionally linked enzymes in a eukaryotic phytoplankton–virus system. We examine arguments for the possible direction of the gene transfer. The virus-to-host direction suggests the existence of ancient viruses that controlled the complex metabolic pathway in order to infect primitive eukaryotic cells. In contrast, the host-to-virus direction suggests that the serial acquisition of genes involved in the same metabolic pathway might have been a strategy for the ancestor of EhVs to stay ahead of their closest relatives in the great evolutionary race for survival. PMID:19451591

  14. Intracellular gene transfer: Reduced hydrophobicity facilitates gene transfer for subunit 2 of cytochrome c oxidase

    PubMed Central

    Daley, Daniel O.; Clifton, Rachel; Whelan, James

    2002-01-01

    Subunit 2 of cytochrome c oxidase (Cox2) in legumes offers a rare opportunity to investigate factors necessary for successful gene transfer of a hydrophobic protein that is usually mitochondrial-encoded. We found that changes in local hydrophobicity were necessary to allow import of this nuclear-encoded protein into mitochondria. All legume species containing both a mitochondrial and nuclear encoded Cox2 displayed a similar pattern, with a large decrease in hydrophobicity evident in the first transmembrane region of the nuclear encoded protein compared with the organelle-encoded protein. Mitochondrial-encoded Cox2 could not be imported into mitochondria under the direction of the mitochondrial targeting sequence that readily supports the import of nuclear encoded Cox2. Removal of the first transmembrane region promotes import ability of the mitochondrial-encoded Cox2. Changing just two amino acids in the first transmembrane region of mitochondrial-encoded Cox2 to the corresponding amino acids in the nuclear encoded Cox2 also promotes import ability, whereas changing the same two amino acids in the nuclear encoded Cox2 to what they are in the mitochondrial-encoded copy prevents import. Therefore, changes in amino acids in the mature protein were necessary and sufficient for gene transfer to allow import under the direction of an appropriate signal to achieve the functional topology of Cox2. PMID:12142462

  15. Systemic Gene Transfer of a Hexosaminidase Variant Using an scAAV9.47 Vector Corrects GM2 Gangliosidosis in Sandhoff Mice.

    PubMed

    Osmon, Karlaina J L; Woodley, Evan; Thompson, Patrick; Ong, Katalina; Karumuthil-Melethil, Subha; Keimel, John G; Mark, Brian L; Mahuran, Don; Gray, Steven J; Walia, Jagdeep S

    2016-07-01

    GM2 gangliosidosis is a group of neurodegenerative diseases caused by β-hexosaminidase A (HexA) enzyme deficiency. There is currently no cure. HexA is composed of two similar, nonidentical subunits, α and β, which must interact with the GM2 activator protein (GM2AP), a substrate-specific cofactor, to hydrolyze GM2 ganglioside. Mutations in either subunit or the activator can result in the accumulation of GM2 ganglioside within neurons throughout the central nervous system. The resulting neuronal cell death induces the primary symptoms of the disease: motor impairment, seizures, and sensory impairments. This study assesses the long-term effects of gene transfer in a Sandhoff (β-subunit knockout) mouse model. The study utilized a modified human β-hexosaminidase α-subunit (μ-subunit) that contains critical sequences from the β-subunit that enables formation of a stable homodimer (HexM) and interaction with GM2AP to hydrolyze GM2 ganglioside. We investigated a self-complementary adeno-associated viral (scAAV) vector expressing HexM, through intravenous injections of the neonatal mice. We monitored one cohort for 8 weeks and another cohort long-term for survival benefit, behavioral, biochemical, and molecular analyses. Untreated Sandhoff disease (SD) control mice reached a humane endpoint at approximately 15 weeks, whereas treated mice had a median survival age of 40 weeks, an approximate 2.5-fold survival advantage. On behavioral tests, the treated mice outperformed their knockout age-matched controls and perform similarly to the heterozygous controls. Through the enzymatic and GM2 ganglioside analyses, we observed a significant decrease in the GM2 ganglioside level, even though the enzyme levels were not significantly increased. Molecular analyses revealed a global distribution of the vector between brain and spinal cord regions. In conclusion, the neonatal delivery of a novel viral vector expressing the human HexM enzyme is effective in ameliorating the SD

  16. BC Transfer System: New Members Policy

    ERIC Educational Resources Information Center

    British Columbia Council on Admissions and Transfer, 2009

    2009-01-01

    The mandate of the British Columbia Council on Admissions and Transfer (BCCAT) includes the responsibility to manage and coordinate the British Columbia (BC) Transfer System. Upon its inception in 1989, the Council inherited a Transfer System that included all BC public institutions, Yukon College, and three private institutions. Since then, new…

  17. An Update on the Learning Transfer System

    ERIC Educational Resources Information Center

    Choi, Myungweon; Ruona, Wendy E. A.

    2008-01-01

    Learning transfer in organizations is a central issue in HRD. Much of the research of the 1980-1990's informed the development of the learning transfer system inventory (Holton, Bates, & Ruona, 2000). However, it's vitally important to continually enhance our understanding of the learning transfer system. In this paper, we reviewed the new…

  18. Ultrasound -Assisted Gene Transfer to Adipose Tissue-Derived Stem/Progenitor Cells (ASCs)

    NASA Astrophysics Data System (ADS)

    Miyamoto, Yoshitaka; Ueno, Hitomi; Hokari, Rei; Yuan, Wenji; Kuno, Shuichi; Kakimoto, Takashi; Enosawa, Shin; Negishi, Yoichi; Yoshinaka, Kiyoshi; Matsumoto, Yoichiro; Chiba, Toshio; Hayashi, Shuji

    2011-09-01

    In recent years, multilineage adipose tissue-derived stem cells (ASCs) have become increasingly attractive as a promising source for cell transplantation and regenerative medicine. Particular interest has been expressed in the potential to make tissue stem cells, such as ASCs and marrow stromal cells (MSCs), differentiate by gene transfection. Gene transfection using highly efficient viral vectors such as adeno- and sendai viruses have been developed for this purpose. Sonoporation, or ultrasound (US)-assisted gene transfer, is an alternative gene manipulation technique which employs the creation of a jet stream by ultrasonic microbubble cavitation. Sonoporation using non-viral vectors is expected to be a much safer, although less efficient, tool for prospective clinical gene therapy. In this report, we assessed the efficacy of the sonoporation technique for gene transfer to ASCs. We isolated and cultured adipocyets from mouse adipose tissue. ASCs that have the potential to differentiate with transformation into adipocytes or osteoblasts were obtained. Using the US-assisted system, plasmid DNA containing beta-galactosidase (beta-Gal) and green fluorescent protein (GFP) genes were transferred to the ASCs. For this purpose, a Sonopore 4000 (NEPAGENE Co.) and a Sonazoid (Daiichi Sankyo Co.) instrument were used in combination. ASCs were subjected to US (3.1 MHz, 50% duty cycle, burst rate 2.0 Hz, intensity 1.2 W/cm2, exposure time 30 sec). We observed that the gene was more efficiently transferred with increased concentrations of plasmid DNA (5-150 μg/mL). However, further optimization of the US parameters is required, as the gene transfer efficiency was still relatively low. In conclusion, we herein demonstrate that a gene can be transferred to ASCs using our US-assisted system. In regenerative medicine, this system might resolve the current issues surrounding the use of viral vectors for gene transfer.

  19. Environmental factors influencing gene transfer agent (GTA) mediated transduction in the subtropical ocean.

    PubMed

    McDaniel, Lauren D; Young, Elizabeth C; Ritchie, Kimberly B; Paul, John H

    2012-01-01

    Microbial genomic sequence analyses have indicated widespread horizontal gene transfer (HGT). However, an adequate mechanism accounting for the ubiquity of HGT has been lacking. Recently, high frequencies of interspecific gene transfer have been documented, catalyzed by Gene Transfer Agents (GTAs) of marine α-Proteobacteria. It has been proposed that the presence of bacterial genes in highly purified viral metagenomes may be due to GTAs. However, factors influencing GTA-mediated gene transfer in the environment have not yet been determined. Several genomically sequenced strains containing complete GTA sequences similar to Rhodobacter capsulatus (RcGTA, type strain) were screened to ascertain if they produced putative GTAs, and at what abundance. Five of nine marine strains screened to date spontaneously produced virus-like particles (VLP's) in stationary phase. Three of these strains have demonstrated gene transfer activity, two of which were documented by this lab. These two strains Roseovarius nubinhibens ISM and Nitratireductor 44B9s, were utilized to produce GTAs designated RnGTA and NrGTA and gene transfer activity was verified in culture. Cell-free preparations of purified RnGTA and NrGTA particles from marked donor strains were incubated with natural microbial assemblages to determine the level of GTA-mediated gene transfer. In conjunction, several ambient environmental parameters were measured including lysogeny indicated by prophage induction. GTA production in culture systems indicated that approximately half of the strains produced GTA-like particles and maximal GTA counts ranged from 10-30% of host abundance. Modeling of GTA-mediated gene transfer frequencies in natural samples, along with other measured environmental variables, indicated a strong relationship between GTA mediated gene transfer and the combined factors of salinity, multiplicity of infection (MOI) and ambient bacterial abundance. These results indicate that GTA-mediated HGT in the

  20. Intrapleural 'outside-in' gene therapy: therapeutics for organs of the chest via gene transfer to the pleura.

    PubMed

    Heguy, Adriana; Crystal, Ronald G

    2005-10-01

    The pleural space is an attractive site for using viral vectors to deliver gene products to the lung parenchyma, other thoracic structures and the systemic circulation. The advantages of intrapleural gene transfer using viral vectors include: (i) easy accessibility; (ii) large surface area; (iii) ability to provide high concentrations of secreted gene products to chest structures; (iv) low risk of detrimental effects of possible vector-induced inflammation compared with intravascular delivery; and (v) because it is local, lower vector doses can be used to deliver therapeutic genes to thoracic structures than less efficient systemic routes. Examples of pleural gene transfer include the use of adenovirus vectors to treat mesothelioma by transiently expressing genes that encode toxic proteins, immunomodulatory molecules or anti-angiogenesis factors. Intrapleural delivery of adeno-associated viral vectors represents an efficient strategy to treat alpha1-antitrypsin (alpha1AT) deficiency, achieving high lung and systemic therapeutic levels of alpha1AT. Intrapleural delivery of gene transfer vectors holds promise for the treatment of diseases requiring transient, localized gene expression, as well as sustained expression of genes to correct hereditary disorders requiring localized or systemic expression of the therapeutic protein.

  1. Transfer function characteristics of super resolving systems

    NASA Technical Reports Server (NTRS)

    Milster, Tom D.; Curtis, Craig H.

    1992-01-01

    Signal quality in an optical storage device greatly depends on the optical system transfer function used to write and read data patterns. The problem is similar to analysis of scanning optical microscopes. Hopkins and Braat have analyzed write-once-read-many (WORM) optical data storage devices. Herein, transfer function analysis of magnetooptic (MO) data storage devices is discussed with respect to improving transfer-function characteristics. Several authors have described improving the transfer function as super resolution. However, none have thoroughly analyzed the MO optical system and effects of the medium. Both the optical system transfer function and effects of the medium of this development are discussed.

  2. Dry Transfer Systems for Used Nuclear Fuel

    SciTech Connect

    Brett W. Carlsen; Michaele BradyRaap

    2012-05-01

    The potential need for a dry transfer system (DTS) to enable retrieval of used nuclear fuel (UNF) for inspection or repackaging will increase as the duration and quantity of fuel in dry storage increases. This report explores the uses for a DTS, identifies associated general functional requirements, and reviews existing and proposed systems that currently perform dry fuel transfers. The focus of this paper is on the need for a DTS to enable transfer of bare fuel assemblies. Dry transfer systems for UNF canisters are currently available and in use for transferring loaded canisters between the drying station and storage and transportation casks.

  3. Genomic analysis of Xanthomonas translucens pathogenic on wheat and barley reveals cross-kingdom gene transfer events and diverse protein delivery systems.

    PubMed

    Gardiner, Donald M; Upadhyaya, Narayana M; Stiller, Jiri; Ellis, Jeff G; Dodds, Peter N; Kazan, Kemal; Manners, John M

    2014-01-01

    In comparison to dicot-infecting bacteria, only limited numbers of genome sequences are available for monocot-infecting and in particular cereal-infecting bacteria. Herein we report the characterisation and genome sequence of Xanthomonas translucens isolate DAR61454 pathogenic on wheat and barley. Based on phylogenetic analysis of the ATP synthase beta subunit (atpD) gene, DAR61454 is most closely related to other X. translucens strains and the sugarcane- and banana- infecting Xanthomonas strains, but shares a type III secretion system (T3SS) with X. translucens pv. graminis and more distantly related xanthomonads. Assays with an adenylate cyclase reporter protein demonstrate that DAR61454's T3SS is functional in delivering proteins to wheat cells. X. translucens DAR61454 also encodes two type VI secretion systems with one most closely related to those found in some strains of the rice infecting strain X. oryzae pv. oryzae but not other xanthomonads. Comparative analysis of 18 different Xanthomonas isolates revealed 84 proteins unique to cereal (i.e. rice) infecting isolates and the wheat/barley infecting DAR61454. Genes encoding 60 of these proteins are found in gene clusters in the X. translucens DAR61454 genome, suggesting cereal-specific pathogenicity islands. However, none of the cereal pathogen specific proteins were homologous to known Xanthomonas spp. effectors. Comparative analysis outside of the bacterial kingdom revealed a nucleoside triphosphate pyrophosphohydrolase encoding gene in DAR61454 also present in other bacteria as well as a number of pathogenic Fusarium species, suggesting that this gene may have been transmitted horizontally from bacteria to the Fusarium lineage of pathogenic fungi. This example further highlights the importance of horizontal gene acquisition from bacteria in the evolution of fungi.

  4. Global Analysis of Horizontal Gene Transfer in Fusarium verticillioides

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The co-occurrence of microbes within plants and other specialized niches may facilitate horizontal gene transfer (HGT) affecting host-pathogen interactions. We recently identified fungal-to-fungal HGTs involving metabolic gene clusters. For a global analysis of HGTs in the maize pathogen Fusarium ve...

  5. Evolution of and horizontal gene transfer in the Endornavirus genus.

    PubMed

    Song, Dami; Cho, Won Kyong; Park, Sang-Ho; Jo, Yeonhwa; Kim, Kook-Hyung

    2013-01-01

    The transfer of genetic information between unrelated species is referred to as horizontal gene transfer. Previous studies have demonstrated that both retroviral and non-retroviral sequences have been integrated into eukaryotic genomes. Recently, we identified many non-retroviral sequences in plant genomes. In this study, we investigated the evolutionary origin and gene transfer of domains present in endornaviruses which are double-stranded RNA viruses. Using the available sequences for endornaviruses, we found that Bell pepper endornavirus-like sequences homologous to the glycosyltransferase 28 domain are present in plants, fungi, and bacteria. The phylogenetic analysis revealed the glycosyltransferase 28 domain of Bell pepper endornavirus may have originated from bacteria. In addition, two domains of Oryza sativa endornavirus, a glycosyltransferase sugar-binding domain and a capsular polysaccharide synthesis protein, also exhibited high similarity to those of bacteria. We found evidence that at least four independent horizontal gene transfer events for the glycosyltransferase 28 domain have occurred among plants, fungi, and bacteria. The glycosyltransferase sugar-binding domains of two proteobacteria may have been horizontally transferred to the genome of Thalassiosira pseudonana. Our study is the first to show that three glycome-related viral genes in the genus Endornavirus have been acquired from marine bacteria by horizontal gene transfer.

  6. Evolution of and Horizontal Gene Transfer in the Endornavirus Genus

    PubMed Central

    Park, Sang-Ho; Jo, Yeonhwa; Kim, Kook-Hyung

    2013-01-01

    The transfer of genetic information between unrelated species is referred to as horizontal gene transfer. Previous studies have demonstrated that both retroviral and non-retroviral sequences have been integrated into eukaryotic genomes. Recently, we identified many non-retroviral sequences in plant genomes. In this study, we investigated the evolutionary origin and gene transfer of domains present in endornaviruses which are double-stranded RNA viruses. Using the available sequences for endornaviruses, we found that Bell pepper endornavirus-like sequences homologous to the glycosyltransferase 28 domain are present in plants, fungi, and bacteria. The phylogenetic analysis revealed the glycosyltransferase 28 domain of Bell pepper endornavirus may have originated from bacteria. In addition, two domains of Oryza sativa endornavirus, a glycosyltransferase sugar-binding domain and a capsular polysaccharide synthesis protein, also exhibited high similarity to those of bacteria. We found evidence that at least four independent horizontal gene transfer events for the glycosyltransferase 28 domain have occurred among plants, fungi, and bacteria. The glycosyltransferase sugar-binding domains of two proteobacteria may have been horizontally transferred to the genome of Thalassiosira pseudonana. Our study is the first to show that three glycome-related viral genes in the genus Endornavirus have been acquired from marine bacteria by horizontal gene transfer. PMID:23667703

  7. Gene transfer agents: phage-like elements of genetic exchange

    PubMed Central

    Lang, Andrew S.; Zhaxybayeva, Olga; Beatty, J. Thomas

    2013-01-01

    Horizontal gene transfer is important in the evolution of bacterial and archaeal genomes. An interesting genetic exchange process is carried out by diverse phage-like gene transfer agents (GTAs) that are found in a wide range of prokaryotes. Although GTAs resemble phages, they lack the hallmark capabilities that define typical phages, and they package random pieces of the producing cell’s genome. In this Review, we discuss the defining characteristics of the GTAs that have been identified to date, along with potential functions for these agents and the possible evolutionary forces that act on the genes involved in their production. PMID:22683880

  8. Evolution of the cutinase gene family: evidence for lateral gene transfer of a candidate Phytophthora virulence factor.

    PubMed

    Belbahri, Lassaad; Calmin, Gautier; Mauch, Felix; Andersson, Jan O

    2008-01-31

    Lateral gene transfer (LGT) can facilitate the acquisition of new functions in recipient lineages, which may enable them to colonize new environments. Several recent publications have shown that gene transfer between prokaryotes and eukaryotes occurs with appreciable frequency. Here we present a study of interdomain gene transfer of cutinases -- well documented virulence factors in fungi -- between eukaryotic plant pathogens Phytophthora species and prokaryotic bacterial lineages. Two putative cutinase genes were cloned from Phytophthora brassicae and Northern blotting experiments showed that these genes are expressed early during the infection of the host Arabidopsis thaliana and induced during cyst germination of the pathogen. Analysis of the gene organisation of this gene family in Phytophthora ramorum and P. sojae showed three and ten copies in tight succession within a region of 5 and 25 kb, respectively, probably indicating a recent expansion in Phytophthora lineages by gene duplications. Bioinformatic analyses identified orthologues only in three genera of Actinobacteria, and in two distantly related eukaryotic groups: oomycetes and fungi. Together with phylogenetic analyses this limited distribution of the gene in the tree of life strongly support a scenario where cutinase genes originated after the origin of land plants in a microbial lineage living in proximity of plants and subsequently were transferred between distantly related plant-degrading microbes. More precisely, a cutinase gene was likely acquired by an ancestor of P. brassicae, P. sojae, P. infestans and P. ramorum, possibly from an actinobacterial source, suggesting that gene transfer might be an important mechanism in the evolution of their virulence. These findings could indeed provide an interesting model system to study acquisition of virulence factors in these important plant pathogens.

  9. High frequency of horizontal gene transfer in the oceans.

    PubMed

    McDaniel, Lauren D; Young, Elizabeth; Delaney, Jennifer; Ruhnau, Fabian; Ritchie, Kim B; Paul, John H

    2010-10-01

    Oceanic bacteria perform many environmental functions, including biogeochemical cycling of many elements, metabolizing of greenhouse gases, functioning in oceanic food webs (microbial loop), and producing valuable natural products and viruses. We demonstrate that the widespread capability of marine bacteria to participate in horizontal gene transfer (HGT) in coastal and oceanic environments may be the result of gene transfer agents (GTAs), viral-like particles produced by α-Proteobacteria. We documented GTA-mediated gene transfer frequencies a thousand to a hundred million times higher than prior estimates of HGT in the oceans, with as high as 47% of the culturable natural microbial community confirmed as gene recipients. These findings suggest a plausible mechanism by which marine bacteria acquire novel traits, thus ensuring resilience in the face of environmental change.

  10. Characterization of Two Cysteine Transfer RNA Genes from Xenopus Laevis

    DTIC Science & Technology

    1984-07-12

    author hereby certifies that the use of any copyrighted material in the dissertation manuscript entitled: "Characterization of two cysteine tRNA genes...Uniformed Services University of the Health Sciences 11 ABSTRACT Title of Thesis: Characterization of Two Cysteine Transfer RNA Genes from Xenopus...method after constructing a set of deletions and reclonlng into the plasmid pUC 8. The DNA fragment is 1737 bp long and contains two cysteine tRNA genes

  11. Fabrication of DNA-antibody-apatite composite layers for cell-targeted gene transfer

    NASA Astrophysics Data System (ADS)

    Yazaki, Yushin; Oyane, Ayako; Araki, Hiroko; Sogo, Yu; Ito, Atsuo; Yamazaki, Atsushi; Tsurushima, Hideo

    2012-12-01

    Surface-mediated gene transfer systems using apatite (Ap)-based composite layers have received increased attention in tissue engineering applications owing to their safety, biocompatibility and relatively high efficiency. In this study, DNA-antibody-apatite composite layers (DA-Ap layers), in which DNA and antibody molecules are immobilized within a matrix of apatite nanocrystals, were fabricated using a biomimetic coating process. They were then assayed for their gene transfer capability for application in a specific cell-targeted gene transfer. A DA-Ap layer that was fabricated with an anti-CD49f antibody showed a higher gene transfer capability to the CD49f-positive CHO-K1 cells than a DNA-apatite composite layer (D-Ap layer). The antibody facilitated the gene transfer capability of the DA-Ap layer only to the specific cells that were expressing corresponding antigens. When the DA-Ap layer was fabricated with an anti-N-cadherin antibody, a higher gene transfer capability compared with the D-Ap layer was found in the N-cadherin-positive P19CL6 cells, but not in the N-cadherin-negative UV♀2 cells or in the P19CL6 cells that were pre-blocked with anti-N-cadherin. Therefore, the antigen-antibody binding that takes place at the cell-layer interface should be responsible for the higher gene transfer capability of the DA-Ap than D-Ap layer. These results suggest that the DA-Ap layer works as a mediator in a specific cell-targeted gene transfer system.

  12. Fabrication of DNA-antibody-apatite composite layers for cell-targeted gene transfer.

    PubMed

    Yazaki, Yushin; Oyane, Ayako; Araki, Hiroko; Sogo, Yu; Ito, Atsuo; Yamazaki, Atsushi; Tsurushima, Hideo

    2012-12-01

    Surface-mediated gene transfer systems using apatite (Ap)-based composite layers have received increased attention in tissue engineering applications owing to their safety, biocompatibility and relatively high efficiency. In this study, DNA-antibody-apatite composite layers (DA-Ap layers), in which DNA and antibody molecules are immobilized within a matrix of apatite nanocrystals, were fabricated using a biomimetic coating process. They were then assayed for their gene transfer capability for application in a specific cell-targeted gene transfer. A DA-Ap layer that was fabricated with an anti-CD49f antibody showed a higher gene transfer capability to the CD49f-positive CHO-K1 cells than a DNA-apatite composite layer (D-Ap layer). The antibody facilitated the gene transfer capability of the DA-Ap layer only to the specific cells that were expressing corresponding antigens. When the DA-Ap layer was fabricated with an anti-N-cadherin antibody, a higher gene transfer capability compared with the D-Ap layer was found in the N-cadherin-positive P19CL6 cells, but not in the N-cadherin-negative UV♀2 cells or in the P19CL6 cells that were pre-blocked with anti-N-cadherin. Therefore, the antigen-antibody binding that takes place at the cell-layer interface should be responsible for the higher gene transfer capability of the DA-Ap than D-Ap layer. These results suggest that the DA-Ap layer works as a mediator in a specific cell-targeted gene transfer system.

  13. Gene transfer into older chicken embryos by ex ovo electroporation.

    PubMed

    Luo, Jiankai; Yan, Xin; Lin, Juntang; Rolfs, Arndt

    2012-07-27

    The chicken embryo provides an excellent model system for studying gene function and regulation during embryonic development. In ovo electroporation is a powerful method to over-express exogenous genes or down-regulate endogenous genes in vivo in chicken embryos(1). Different structures such as DNA plasmids encoding genes(2-4), small interfering RNA (siRNA) plasmids(5), small synthetic RNA oligos(6), and morpholino antisense oligonucleotides(7) can be easily transfected into chicken embryos by electroporation. However, the application of in ovo electroporation is limited to embryos at early incubation stages (younger than stage HH20--according to Hamburg and Hamilton)(8) and there are some disadvantages for its application in embryos at later stages (older than stage HH22--approximately 3.5 days of development). For example, the vitelline membrane at later stages is usually stuck to the shall membrane and opening a window in the shell causes rupture of the vessels, resulting in death of the embryos; older embryos are covered by vitelline and allantoic vessels, where it is difficult to access and manipulate the embryos; older embryos move vigorously and is difficult to control the orientation through a relatively small window in the shell. In this protocol we demonstrate an ex ovo electroporation method for gene transfer into chicken embryos at late stages (older than stage HH22). For ex ovo electroporation, embryos are cultured in Petri dishes(9) and the vitelline and allantoic vessels are widely spread. Under these conditions, the older chicken embryos are easily accessed and manipulated. Therefore, this method overcomes the disadvantages of in ovo electroporation applied to the older chicken embryos. Using this method, plasmids can be easily transfected into different parts of the older chicken embryos(10-12).

  14. Lateral Transfer of Genes and Gene Fragments in Staphylococcus Extends beyond Mobile Elements ▿ †

    PubMed Central

    Chan, Cheong Xin; Beiko, Robert G.; Ragan, Mark A.

    2011-01-01

    The widespread presence of antibiotic resistance and virulence among Staphylococcus isolates has been attributed in part to lateral genetic transfer (LGT), but little is known about the broader extent of LGT within this genus. Here we report the first systematic study of the modularity of genetic transfer among 13 Staphylococcus genomes covering four distinct named species. Using a topology-based phylogenetic approach, we found, among 1,354 sets of homologous genes examined, strong evidence of LGT in 368 (27.1%) gene sets, and weaker evidence in another 259 (19.1%). Within-gene and whole-gene transfer contribute almost equally to the topological discordance of these gene sets against a reference phylogeny. Comparing genetic transfer in single-copy and in multicopy gene sets, we observed a higher frequency of LGT in the latter, and a substantial functional bias in cases of whole-gene transfer (little such bias was observed in cases of fragmentary genetic transfer). We found evidence that lateral transfer, particularly of entire genes, impacts not only functions related to antibiotic, drug, and heavy-metal resistance, as well as membrane transport, but also core informational and metabolic functions not associated with mobile elements. Although patterns of sequence similarity support the cohesion of recognized species, LGT within S. aureus appears frequently to disrupt clonal complexes. Our results demonstrate that LGT and gene duplication play important parts in functional innovation in staphylococcal genomes. PMID:21622749

  15. Immunomodulation by mucosal gene transfer using TGF-beta DNA.

    PubMed Central

    Kuklin, N A; Daheshia, M; Chun, S; Rouse, B T

    1998-01-01

    This report evaluates the efficacy of DNA encoding TGF-beta administered mucosally to suppress immunity and modulate the immunoinflammatory response to herpes simplex virus (HSV) infection. A single intranasal administration of an eukaryotic expression vector encoding TGF-beta1 led to expression in the lung and lymphoid tissue. T cell-mediated immune responses to HSV infection were suppressed with this effect persisting as measured by the delayed-type hypersensitivity reaction for at least 7 wk. Treated animals were more susceptible to systemic infection with HSV. Multiple prophylactic mucosal administrations of TGF-beta DNA also suppressed the severity of ocular lesions caused by HSV infection, although no effects on this immunoinflammatory response were evident after therapeutic treatment with TGF-beta DNA. Our results demonstrate that the direct mucosal gene transfer of immunomodulatory cytokines provides a convenient means of modulating immunity and influencing the expression of inflammatory disorders. PMID:9664086

  16. Horizontal functional gene transfer from bacteria to fishes

    PubMed Central

    Sun, Bao-Fa; Li, Tong; Xiao, Jin-Hua; Jia, Ling-Yi; Liu, Li; Zhang, Peng; Murphy, Robert W.; He, Shun-Min; Huang, Da-Wei

    2015-01-01

    Invertebrates can acquire functional genes via horizontal gene transfer (HGT) from bacteria but fishes are not known to do so. We provide the first reliable evidence of one HGT event from marine bacteria to fishes. The HGT appears to have occurred after emergence of the teleosts. The transferred gene is expressed and regulated developmentally. Its successful integration and expression may change the genetic and metabolic repertoire of fishes. In addition, this gene contains conserved domains and similar tertiary structures in fishes and their putative donor bacteria. Thus, it may function similarly in both groups. Evolutionary analyses indicate that it evolved under purifying selection, further indicating its conserved function. We document the first likely case of HGT of functional gene from prokaryote to fishes. This discovery certifies that HGT can influence vertebrate evolution. PMID:26691285

  17. Horizontal functional gene transfer from bacteria to fishes.

    PubMed

    Sun, Bao-Fa; Li, Tong; Xiao, Jin-Hua; Jia, Ling-Yi; Liu, Li; Zhang, Peng; Murphy, Robert W; He, Shun-Min; Huang, Da-Wei

    2015-12-22

    Invertebrates can acquire functional genes via horizontal gene transfer (HGT) from bacteria but fishes are not known to do so. We provide the first reliable evidence of one HGT event from marine bacteria to fishes. The HGT appears to have occurred after emergence of the teleosts. The transferred gene is expressed and regulated developmentally. Its successful integration and expression may change the genetic and metabolic repertoire of fishes. In addition, this gene contains conserved domains and similar tertiary structures in fishes and their putative donor bacteria. Thus, it may function similarly in both groups. Evolutionary analyses indicate that it evolved under purifying selection, further indicating its conserved function. We document the first likely case of HGT of functional gene from prokaryote to fishes. This discovery certifies that HGT can influence vertebrate evolution.

  18. Recent events dominate interdomain lateral gene transfers between prokaryotes and eukaryotes and, with the exception of endosymbiotic gene transfers, few ancient transfer events persist

    PubMed Central

    Katz, Laura A.

    2015-01-01

    While there is compelling evidence for the impact of endosymbiotic gene transfer (EGT; transfer from either mitochondrion or chloroplast to the nucleus) on genome evolution in eukaryotes, the role of interdomain transfer from bacteria and/or archaea (i.e. prokaryotes) is less clear. Lateral gene transfers (LGTs) have been argued to be potential sources of phylogenetic information, particularly for reconstructing deep nodes that are difficult to recover with traditional phylogenetic methods. We sought to identify interdomain LGTs by using a phylogenomic pipeline that generated 13 465 single gene trees and included up to 487 eukaryotes, 303 bacteria and 118 archaea. Our goals include searching for LGTs that unite major eukaryotic clades, and describing the relative contributions of LGT and EGT across the eukaryotic tree of life. Given the difficulties in interpreting single gene trees that aim to capture the approximately 1.8 billion years of eukaryotic evolution, we focus on presence–absence data to identify interdomain transfer events. Specifically, we identify 1138 genes found only in prokaryotes and representatives of three or fewer major clades of eukaryotes (e.g. Amoebozoa, Archaeplastida, Excavata, Opisthokonta, SAR and orphan lineages). The majority of these genes have phylogenetic patterns that are consistent with recent interdomain LGTs and, with the notable exception of EGTs involving photosynthetic eukaryotes, we detect few ancient interdomain LGTs. These analyses suggest that LGTs have probably occurred throughout the history of eukaryotes, but that ancient events are not maintained unless they are associated with endosymbiotic gene transfer among photosynthetic lineages. PMID:26323756

  19. Lunox storage and transfer system

    NASA Technical Reports Server (NTRS)

    1987-01-01

    This semester, efforts were concentrated on the design of the Lunox transfer line from the storage area to the launch site. Emphasis was placed on flow and heat transfer problems and their remedies by reducing the effect of radiation by selecting materials for storage tanks, transfer lines and insulation. The design for the storage tank was based on a medium sized Lunox production facility of 6,000 metric tons per year and the frequency of transportation of Lunox from lunar launch site to lower lunar orbit of four launches per month. The design included the selection of materials for cryogenic storage, insulation and radiation shielding. Lunox was pumped to the storage area near the launch site through a piping network designed for maximum mass flow rate with a minimum boil off. The entire network incorporated specially designed radiation shields made of material which was lightweight and low in secondary radiation.

  20. The interconnection between biofilm formation and horizontal gene transfer.

    PubMed

    Madsen, Jonas Stenløkke; Burmølle, Mette; Hansen, Lars Hestbjerg; Sørensen, Søren Johannes

    2012-07-01

    Recent research has revealed that horizontal gene transfer and biofilm formation are connected processes. Although published research investigating this interconnectedness is still limited, we will review this subject in order to highlight the potential of these observations because of their believed importance in the understanding of the adaptation and subsequent evolution of social traits in bacteria. Here, we discuss current evidence for such interconnectedness centred on plasmids. Horizontal transfer rates are typically higher in biofilm communities compared with those in planktonic states. Biofilms, furthermore, promote plasmid stability and may enhance the host range of mobile genetic elements that are transferred horizontally. Plasmids, on the other hand, are very well suited to promote the evolution of social traits such as biofilm formation. This, essentially, transpires because plasmids are independent replicons that enhance their own success by promoting inter-bacterial interactions. They typically also carry genes that heighten their hosts' direct fitness. Furthermore, current research shows that the so-called mafia traits encoded on mobile genetic elements can enforce bacteria to maintain stable social interactions. It also indicates that horizontal gene transfer ultimately enhances the relatedness of bacteria carrying the mobile genetic elements of the same origin. The perspective of this review extends to an overall interconnectedness between horizontal gene transfer, mobile genetic elements and social evolution of bacteria.

  1. [Gene doping: gene transfer and possible molecular detection].

    PubMed

    Argüelles, Carlos Francisco; Hernández-Zamora, Edgar

    2007-01-01

    The use of illegal substances in sports to enhance athletic performance during competition has caused international sports organizations such as the COI and WADA to take anti doping measures. A new doping method know as gene doping is defined as "the non-therapeutic use of genes, genetic elements and/or cells that have the capacity to enhance athletic performance". However, gene doping in sports is not easily identified and can cause serious consequences. Molecular biology techniques are needed in order to distinguish the difference between a "normal" and an "altered" genome. Further, we need to develop new analytic methods and biological molecular techniques in anti-doping laboratories, and design programs that avoid the non therapeutic use of genes.

  2. Saturn facility oil transfer automation system

    SciTech Connect

    Joseph, Nathan R.; Thomas, Rayburn Dean; Lewis, Barbara Ann; Malagon, Hector M.

    2014-02-01

    The Saturn accelerator, owned by Sandia National Laboratories, has been in operation since the early 1980s and still has many of the original systems. A critical legacy system is the oil transfer system which transfers 250,000 gallons of transformer oil from outside storage tanks to the Saturn facility. The oil transfer system was iden- ti ed for upgrade to current technology standards. Using the existing valves, pumps, and relay controls, the system was automated using the National Instruments cRIO FGPA platform. Engineered safety practices, including a failure mode e ects analysis, were used to develop error handling requirements. The uniqueness of the Saturn Oil Automated Transfer System (SOATS) is in the graphical user interface. The SOATS uses an HTML interface to communicate to the cRIO, creating a platform independent control system. The SOATS was commissioned in April 2013.

  3. Antibacterial gene transfer across the tree of life

    PubMed Central

    Metcalf, Jason A; Funkhouser-Jones, Lisa J; Brileya, Kristen; Reysenbach, Anna-Louise; Bordenstein, Seth R

    2014-01-01

    Though horizontal gene transfer (HGT) is widespread, genes and taxa experience biased rates of transferability. Curiously, independent transmission of homologous DNA to archaea, bacteria, eukaryotes, and viruses is extremely rare and often defies ecological and functional explanations. Here, we demonstrate that a bacterial lysozyme family integrated independently in all domains of life across diverse environments, generating the only glycosyl hydrolase 25 muramidases in plants and archaea. During coculture of a hydrothermal vent archaeon with a bacterial competitor, muramidase transcription is upregulated. Moreover, recombinant lysozyme exhibits broad-spectrum antibacterial action in a dose-dependent manner. Similar to bacterial transfer of antibiotic resistance genes, transfer of a potent antibacterial gene across the universal tree seemingly bestows a niche-transcending adaptation that trumps the barriers against parallel HGT to all domains. The discoveries also comprise the first characterization of an antibacterial gene in archaea and support the pursuit of antibiotics in this underexplored group. DOI: http://dx.doi.org/10.7554/eLife.04266.001 PMID:25422936

  4. Replacing and Additive Horizontal Gene Transfer in Streptococcus

    PubMed Central

    Choi, Sang Chul; Rasmussen, Matthew D.; Hubisz, Melissa J.; Gronau, Ilan; Stanhope, Michael J.; Siepel, Adam

    2012-01-01

    The prominent role of Horizontal Gene Transfer (HGT) in the evolution of bacteria is now well documented, but few studies have differentiated between evolutionary events that predominantly cause genes in one lineage to be replaced by homologs from another lineage (“replacing HGT”) and events that result in the addition of substantial new genomic material (“additive HGT”). Here in, we make use of the distinct phylogenetic signatures of replacing and additive HGTs in a genome-wide study of the important human pathogen Streptococcus pyogenes (SPY) and its close relatives S. dysgalactiae subspecies equisimilis (SDE) and S. dysgalactiae subspecies dysgalactiae (SDD). Using recently developed statistical models and computational methods, we find evidence for abundant gene flow of both kinds within each of the SPY and SDE clades and of reduced levels of exchange between SPY and SDD. In addition, our analysis strongly supports a pronounced asymmetry in SPY–SDE gene flow, favoring the SPY-to-SDE direction. This finding is of particular interest in light of the recent increase in virulence of pathogenic SDE. We find much stronger evidence for SPY–SDE gene flow among replacing than among additive transfers, suggesting a primary influence from homologous recombination between co-occurring SPY and SDE cells in human hosts. Putative virulence genes are correlated with transfer events, but this correlation is found to be driven by additive, not replacing, HGTs. The genes affected by additive HGTs are enriched for functions having to do with transposition, recombination, and DNA integration, consistent with previous findings, whereas replacing HGTs seen to influence a more diverse set of genes. Additive transfers are also found to be associated with evidence of positive selection. These findings shed new light on the manner in which HGT has shaped pathogenic bacterial genomes. PMID:22617954

  5. Horizontal gene transfer in the human gastrointestinal tract: potential spread of antibiotic resistance genes

    PubMed Central

    Huddleston, Jennifer R

    2014-01-01

    Bacterial infections are becoming increasingly difficult to treat due to widespread antibiotic resistance among pathogens. This review aims to give an overview of the major horizontal transfer mechanisms and their evolution and then demonstrate the human lower gastrointestinal tract as an environment in which horizontal gene transfer of resistance determinants occurs. Finally, implications for antibiotic usage and the development of resistant infections and persistence of antibiotic resistance genes in populations as a result of horizontal gene transfer in the large intestine will be discussed. PMID:25018641

  6. Gene transfer and mutagenesis mediated by Sleeping Beauty transposon in Nile tilapia (Oreochromis niloticus).

    PubMed

    He, Xiaozhen; Li, Jie; Long, Yong; Song, Guili; Zhou, Peiyong; Liu, Qiuxiang; Zhu, Zuoyan; Cui, Zongbin

    2013-10-01

    The success of gene transfer has been demonstrated in many of vertebrate species, whereas the efficiency of producing transgenic animals remains pretty low due to the random integration of foreign genes into a recipient genome. The Sleeping Beauty (SB) transposon is able to improve the efficiency of gene transfer in zebrafish and mouse, but its activity in tilapia (Oreochromis niloticus) has yet to be characterized. Herein, we demonstrate the potential of using the SB transposon system as an effective tool for gene transfer and insertional mutagenesis in tilapia. A transgenic construct pT2/tiHsp70-SB11 was generated by subcloning the promoter of tilapia heat shock protein 70 (tiHsp70) gene, the SB11 transposase gene and the carp β-actin gene polyadenylation signal into the second generation of SB transposon. Transgenic tilapia was produced by microinjection of this construct with in vitro synthesized capped SB11 mRNA. SB11 transposon was detected in 28.89 % of founders, 12.9 % of F1 and 43.75 % of F2. Analysis of genomic sequences flanking integrated transposons indicates that this transgenic tilapia line carries two copies of SB transposon, which landed into two different endogenous genes. Induced expression of SB11 gene after heat shock was detected using reverse transcription PCR in F2 transgenic individuals. In addition, the Cre/loxP system was introduced to delete the SB11 cassette for stabilization of gene interruption and bio-safety. These findings suggest that the SB transposon system is active and can be used for efficient gene transfer and insertional mutagenesis in tilapia.

  7. Horizontal gene transfer in eukaryotes: The weak-link model

    PubMed Central

    Huang, Jinling

    2013-01-01

    The significance of horizontal gene transfer (HGT) in eukaryotic evolution remains controversial. Although many eukaryotic genes are of bacterial origin, they are often interpreted as being derived from mitochondria or plastids. Because of their fixed gene pool and gene loss, however, mitochondria and plastids alone cannot adequately explain the presence of all, or even the majority, of bacterial genes in eukaryotes. Available data indicate that no insurmountable barrier to HGT exists, even in complex multicellular eukaryotes. In addition, the discovery of both recent and ancient HGT events in all major eukaryotic groups suggests that HGT has been a regular occurrence throughout the history of eukaryotic evolution. A model of HGT is proposed that suggests both unicellular and early developmental stages as likely entry points for foreign genes into multicellular eukaryotes. PMID:24037739

  8. Horizontal gene transfer in eukaryotes: the weak-link model.

    PubMed

    Huang, Jinling

    2013-10-01

    The significance of horizontal gene transfer (HGT) in eukaryotic evolution remains controversial. Although many eukaryotic genes are of bacterial origin, they are often interpreted as being derived from mitochondria or plastids. Because of their fixed gene pool and gene loss, however, mitochondria and plastids alone cannot adequately explain the presence of all, or even the majority, of bacterial genes in eukaryotes. Available data indicate that no insurmountable barrier to HGT exists, even in complex multicellular eukaryotes. In addition, the discovery of both recent and ancient HGT events in all major eukaryotic groups suggests that HGT has been a regular occurrence throughout the history of eukaryotic evolution. A model of HGT is proposed that suggests both unicellular and early developmental stages as likely entry points for foreign genes into multicellular eukaryotes.

  9. Bacterial Genes in the Aphid Genome: Absence of Functional Gene Transfer from Buchnera to Its Host

    PubMed Central

    Nikoh, Naruo; McCutcheon, John P.; Kudo, Toshiaki; Miyagishima, Shin-ya; Moran, Nancy A.; Nakabachi, Atsushi

    2010-01-01

    Genome reduction is typical of obligate symbionts. In cellular organelles, this reduction partly reflects transfer of ancestral bacterial genes to the host genome, but little is known about gene transfer in other obligate symbioses. Aphids harbor anciently acquired obligate mutualists, Buchnera aphidicola (Gammaproteobacteria), which have highly reduced genomes (420–650 kb), raising the possibility of gene transfer from ancestral Buchnera to the aphid genome. In addition, aphids often harbor other bacteria that also are potential sources of transferred genes. Previous limited sampling of genes expressed in bacteriocytes, the specialized cells that harbor Buchnera, revealed that aphids acquired at least two genes from bacteria. The newly sequenced genome of the pea aphid, Acyrthosiphon pisum, presents the first opportunity for a complete inventory of genes transferred from bacteria to the host genome in the context of an ancient obligate symbiosis. Computational screening of the entire A. pisum genome, followed by phylogenetic and experimental analyses, provided strong support for the transfer of 12 genes or gene fragments from bacteria to the aphid genome: three LD–carboxypeptidases (LdcA1, LdcA2,ψLdcA), five rare lipoprotein As (RlpA1-5), N-acetylmuramoyl-L-alanine amidase (AmiD), 1,4-beta-N-acetylmuramidase (bLys), DNA polymerase III alpha chain (ψDnaE), and ATP synthase delta chain (ψAtpH). Buchnera was the apparent source of two highly truncated pseudogenes (ψDnaE and ψAtpH). Most other transferred genes were closely related to genes from relatives of Wolbachia (Alphaproteobacteria). At least eight of the transferred genes (LdcA1, AmiD, RlpA1-5, bLys) appear to be functional, and expression of seven (LdcA1, AmiD, RlpA1-5) are highly upregulated in bacteriocytes. The LdcAs and RlpAs appear to have been duplicated after transfer. Our results excluded the hypothesis that genome reduction in Buchnera has been accompanied by gene transfer to the host

  10. Disconnecting the Yin and Yang Relation of Epidermal Growth Factor Receptor (EGFR)-Mediated Delivery: A Fully Synthetic, EGFR-Targeted Gene Transfer System Avoiding Receptor Activation

    PubMed Central

    Schäfer, A.; Pahnke, A.; Schaffert, D.; van Weerden, W.M.; de Ridder, C.M.A.; Rödl, W.; Vetter, A.; Spitzweg, C.; Kraaij, R.; Wagner, E.

    2011-01-01

    Abstract The epidermal growth factor receptor (EGFR) is upregulated within a high percentage of solid tumors and hence is an attractive target for tumor-targeted therapies including gene therapy. The natural EGFR ligand epidermal growth factor (EGF) has been used for this purpose, despite the risk of mitogenic effects due to EGFR activation. We have developed a fully synthetic, EGFR-targeted gene delivery system based on PEGylated linear polyethylenimine (LPEI), allowing evaluation of different EGFR-binding peptides in terms of transfection efficiency and EGFR activation. Peptide sequences directly derived from the human EGF molecule enhanced transfection efficiency with concomitant EGFR activation. Only the EGFR-binding peptide GE11, which has been identified by phage display technique, showed specific enhancement of transfection on EGFR-overexpressing tumor cells including glioblastoma and hepatoma, but without EGFR activation. EGFR targeting led to high levels of cell association of fluorescently labeled polyplexes after only 30 min of incubation. EGF pretreatment of cells induced enhanced cellular internalization of all polyplex types tested, pointing at generally enhanced macropinocytosis. EGF polyplexes diminished cell surface expression of EGFR for up to 4 hr, whereas GE11 polyplexes did not. In a clinically relevant orthotopic prostate cancer model, intratumorally injected GE11 polyplexes were superior in inducing transgene expression when compared with untargeted polyplexes. PMID:21644815

  11. Insight into a reversible energy transfer system.

    PubMed

    Gao, Ming Xuan; Zou, Hong Yan; Gao, Peng Fei; Liu, Yue; Li, Na; Li, Yuan Fang; Huang, Cheng Zhi

    2016-09-15

    Resonance energy transfer (RET) processes have wide applications; these processes involve a unidirectional energy transfer from a particular donor to a particular acceptor. Here, we report a plasmonic resonance energy transfer (PRET), which occurs from the surface of gold nanoparticles to fluorescent organic dyes, and coexists with a nanometal surface energy transfer (NSET) that operates in the reverse direction. The coexistence of both PRET and NSET in opposite directions means that the roles of both donor and acceptor can be interchanged, which could be identified by using spectrofluorometric measurements and light scattering dark field microscopic imaging. The experimental data could be further theoretically supported using Persson and Lang's model, the quasi-static approximation and finite-difference time-domain simulation. Moreover, disruption of the PRET process by altering the energy transfer pairs suggests that interactions occur inside the reversible energy transfer system, which manifest by increasing the fluorescence quenching efficiency of the NSET process.

  12. Antibiotics and gene transfer in swine gut bacteria

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The mammalian gastrointestinal (GI) tract hosts a diverse collection bacteria, most of which are beneficial for host health. This bacterial community also supports a community of viruses that infect bacteria (called bacteriophages or phages). Phages transfer genes between bacteria, and phage-media...

  13. Quasispecies theory for horizontal gene transfer and recombination

    NASA Astrophysics Data System (ADS)

    Muñoz, Enrique; Park, Jeong-Man; Deem, Michael W.

    2008-12-01

    We introduce a generalization of the parallel, or Crow-Kimura, and Eigen models of molecular evolution to represent the exchange of genetic information between individuals in a population. We study the effect of different schemes of genetic recombination on the steady-state mean fitness and distribution of individuals in the population, through an analytic field theoretic mapping. We investigate both horizontal gene transfer from a population and recombination between pairs of individuals. Somewhat surprisingly, these nonlinear generalizations of quasispecies theory to modern biology are analytically solvable. For two-parent recombination, we find two selected phases, one of which is spectrally rigid. We present exact analytical formulas for the equilibrium mean fitness of the population, in terms of a maximum principle, which are generally applicable to any permutation invariant replication rate function. For smooth fitness landscapes, we show that when positive epistatic interactions are present, recombination or horizontal gene transfer introduces a mild load against selection. Conversely, if the fitness landscape exhibits negative epistasis, horizontal gene transfer or recombination introduces an advantage by enhancing selection towards the fittest genotypes. These results prove that the mutational deterministic hypothesis holds for quasispecies models. For the discontinuous single sharp peak fitness landscape, we show that horizontal gene transfer has no effect on the fitness, while recombination decreases the fitness, for both the parallel and the Eigen models. We present numerical and analytical results as well as phase diagrams for the different cases.

  14. Electron transfer and energy transfer through bridged systems. I. Formalism

    NASA Astrophysics Data System (ADS)

    Reimers, J. R.; Hush, N. S.

    1989-07-01

    A time-dependent formalism is developed for reactions in which energy (vibrational or electronic excitation, electron or hole transfer, etc.) is transferred coherently between centres through a bridge. This approach is inspired by the Robinson and Frosch model of energy transfer within two-level systems. This formalism yields a completely general algorithm which, in particular limits, reduces to a generalised form of both Fermi's golden rule and Rabi's rate equation, and, in so doing, unifies many existing theories. It is shown that, only in the limit of the bridge states being non-resonant with the initial and final states, can the full problem be represented by an effective two-level model. Existing methods based upon Löwdin diagonalization are shown to be appropriate only when this limit applies, and ambiguities which arise from the ad hoc nature of these methods are resolved. Also, it is typically only in this limit that the transfer of energy proceeds exponentially in time and can be described by a simple single-parameter rate constant. Only problems which can be modelled using a single set of quantum numbers are treated in this paper. Applications and more general problems are treated in subsequent papers.

  15. Biased gene transfer mimics patterns created through shared ancestry

    PubMed Central

    Andam, Cheryl P.; Williams, David; Gogarten, J. Peter

    2010-01-01

    In phylogenetic reconstruction, two types of bacterial tyrosyl-tRNA synthetases (TyrRS) form distinct clades with many bacterial phyla represented in both clades. Very few taxa possess both forms, and maximum likelihood analysis of the distribution of TyrRS types suggests horizontal gene transfer (HGT), rather than an ancient duplication followed by differential gene loss, as the contributor to the evolutionary history of TyrRS in bacteria. However, for each TyrRS type, phylogenetic reconstruction yields phylogenies similar to the ribosomal phylogeny, revealing that frequent gene transfer has not destroyed the expected phylogeny; rather, the expected phylogenetic signal was reinforced or even created by HGT. We show that biased HGT can mimic patterns created through shared ancestry by in silico simulation. Furthermore, in cases where genomic synteny is sufficient to allow comparisons of relative gene positions, both tyrRS types occupy equivalent positions in closely related genomes, rejecting the loss hypothesis. Although the two types of bacterial TyrRS are only distantly related and only rarely coexist in a single genome, they have many features in common with alleles that are swapped between related lineages. We propose to label these functionally similar homologs as homeoalleles. We conclude that the observed phylogenetic pattern reflects both vertical inheritance and biased HGT and that the signal caused by common organismal descent is difficult to distinguish from the signal due to biased gene transfer. PMID:20495090

  16. Biased gene transfer mimics patterns created through shared ancestry.

    PubMed

    Andam, Cheryl P; Williams, David; Gogarten, J Peter

    2010-06-08

    In phylogenetic reconstruction, two types of bacterial tyrosyl-tRNA synthetases (TyrRS) form distinct clades with many bacterial phyla represented in both clades. Very few taxa possess both forms, and maximum likelihood analysis of the distribution of TyrRS types suggests horizontal gene transfer (HGT), rather than an ancient duplication followed by differential gene loss, as the contributor to the evolutionary history of TyrRS in bacteria. However, for each TyrRS type, phylogenetic reconstruction yields phylogenies similar to the ribosomal phylogeny, revealing that frequent gene transfer has not destroyed the expected phylogeny; rather, the expected phylogenetic signal was reinforced or even created by HGT. We show that biased HGT can mimic patterns created through shared ancestry by in silico simulation. Furthermore, in cases where genomic synteny is sufficient to allow comparisons of relative gene positions, both tyrRS types occupy equivalent positions in closely related genomes, rejecting the loss hypothesis. Although the two types of bacterial TyrRS are only distantly related and only rarely coexist in a single genome, they have many features in common with alleles that are swapped between related lineages. We propose to label these functionally similar homologs as homeoalleles. We conclude that the observed phylogenetic pattern reflects both vertical inheritance and biased HGT and that the signal caused by common organismal descent is difficult to distinguish from the signal due to biased gene transfer.

  17. Optical Energy Transfer and Conversion System

    NASA Technical Reports Server (NTRS)

    Stone, William C. (Inventor); Hogan, Bartholomew P. (Inventor)

    2015-01-01

    An optical power transfer system comprising a fiber spooler, a fiber optic rotary joint mechanically connected to the fiber spooler, and an electrical power extraction subsystem connected to the fiber optic rotary joint with an optical waveguide. Optical energy is generated at and transferred from a base station through fiber wrapped around the spooler, through the rotary joint, and ultimately to the power extraction system at a remote mobility platform for conversion to another form of energy.

  18. Quartet analysis of putative horizontal gene transfer in Crenarchaeota.

    PubMed

    Ching, Travers H; Yoza, Brandon A; Li, Qing X

    2014-02-01

    Horizontal gene transfers (HGT) between four Crenarchaeota species (Metallosphaera cuprina Ar-4T, Acidianus hospitalis W1T, Vulcanisaeta moutnovskia 768-28T, and Pyrobaculum islandicum DSM 4184T) were investigated with quartet analysis. Strong support was found for individual genes that disagree with the phylogeny of the majority, implying genomic mosaicism. One such gene, a ferredoxin-related gene, was investigated further and incorporated into a larger phylogeny, which provided evidence for HGT of this gene from the Vulcanisaeta lineage to the Acidianus lineage. This is the first application of quartet analysis of HGT for the phylum Crenarchaeota. The results have shown that quartet analysis is a powerful technique to screen homologous sequences for putative HGTs and is useful in visually describing genomic mosaicism and HGT within four taxa.

  19. Rescuing the Failing Heart by Targeted Gene Transfer

    PubMed Central

    Kawase, Yoshiaki; Ladage, Dennis; Hajjar, Roger J.

    2011-01-01

    Congestive heart failure is a major cause of morbidity and mortality in the US. While progress in conventional treatments is making steady and incremental gains to reduce heart failure mortality, there is a critical need to explore new therapeutic approaches. Gene therapy was initially applied in the clinical setting for inherited monogenic disorders. It is now apparent that gene therapy has broader potential that also includes acquired polygenic diseases, such as congestive heart failure. Recent advances in understanding of the molecular basis of myocardial dysfunction, together with the evolution of increasingly efficient gene transfer technology, has placed heart failure within reach of gene-based therapy. Furthermore, the recent successful and safe completion of a phase 2 trial targeting the sarcoplasmic reticulum calcium ATPase pump (SERCA2a) along with the start of more recent phase 1 trials usher a new era for gene therapy for the treatment of heart failure. PMID:21371634

  20. Tumor specific ultrasound enhanced gene transfer in vivo with novel liposomal bubbles.

    PubMed

    Suzuki, Ryo; Takizawa, Tomoko; Negishi, Yoichi; Utoguchi, Naoki; Sawamura, Kaori; Tanaka, Kumiko; Namai, Eisuke; Oda, Yusuke; Matsumura, Yasuhiro; Maruyama, Kazuo

    2008-01-22

    Bubble liposomes (liposomes which entrap an ultrasound imaging gas) may constitute a unique system for delivering various molecules efficiently into mammalian cells in vitro. In this study, Bubble liposomes were compared with cationic lipid (CL)-DNA complexes as potential gene delivery carriers into tumor in vivo. The delivery of genes by Bubble liposomes depended on the intensity of the applied ultrasound. Transfection efficiency plateaued at 0.7 W/cm(2) ultrasound intensity. Bubble liposomes efficiently transferred genes into cultured cells even when the cells were exposed to ultrasound for only 1 s. In addition, Bubble liposomes could introduce the luciferase gene more effectively than CL-DNA complexes into mouse ascites tumor cells and solid tumor tissue. We conclude that the combination of Bubble liposomes and ultrasound is a minimally-invasive and tumor specific gene transfer method in vivo.

  1. Endosymbiotic gene transfer from prokaryotic pangenomes: Inherited chimerism in eukaryotes.

    PubMed

    Ku, Chuan; Nelson-Sathi, Shijulal; Roettger, Mayo; Garg, Sriram; Hazkani-Covo, Einat; Martin, William F

    2015-08-18

    Endosymbiotic theory in eukaryotic-cell evolution rests upon a foundation of three cornerstone partners--the plastid (a cyanobacterium), the mitochondrion (a proteobacterium), and its host (an archaeon)--and carries a corollary that, over time, the majority of genes once present in the organelle genomes were relinquished to the chromosomes of the host (endosymbiotic gene transfer). However, notwithstanding eukaryote-specific gene inventions, single-gene phylogenies have never traced eukaryotic genes to three single prokaryotic sources, an issue that hinges crucially upon factors influencing phylogenetic inference. In the age of genomes, single-gene trees, once used to test the predictions of endosymbiotic theory, now spawn new theories that stand to eventually replace endosymbiotic theory with descriptive, gene tree-based variants featuring supernumerary symbionts: prokaryotic partners distinct from the cornerstone trio and whose existence is inferred solely from single-gene trees. We reason that the endosymbiotic ancestors of mitochondria and chloroplasts brought into the eukaryotic--and plant and algal--lineage a genome-sized sample of genes from the proteobacterial and cyanobacterial pangenomes of their respective day and that, even if molecular phylogeny were artifact-free, sampling prokaryotic pangenomes through endosymbiotic gene transfer would lead to inherited chimerism. Recombination in prokaryotes (transduction, conjugation, transformation) differs from recombination in eukaryotes (sex). Prokaryotic recombination leads to pangenomes, and eukaryotic recombination leads to vertical inheritance. Viewed from the perspective of endosymbiotic theory, the critical transition at the eukaryote origin that allowed escape from Muller's ratchet--the origin of eukaryotic recombination, or sex--might have required surprisingly little evolutionary innovation.

  2. Endosymbiotic gene transfer from prokaryotic pangenomes: Inherited chimerism in eukaryotes

    PubMed Central

    Ku, Chuan; Nelson-Sathi, Shijulal; Roettger, Mayo; Garg, Sriram; Hazkani-Covo, Einat; Martin, William F.

    2015-01-01

    Endosymbiotic theory in eukaryotic-cell evolution rests upon a foundation of three cornerstone partners—the plastid (a cyanobacterium), the mitochondrion (a proteobacterium), and its host (an archaeon)—and carries a corollary that, over time, the majority of genes once present in the organelle genomes were relinquished to the chromosomes of the host (endosymbiotic gene transfer). However, notwithstanding eukaryote-specific gene inventions, single-gene phylogenies have never traced eukaryotic genes to three single prokaryotic sources, an issue that hinges crucially upon factors influencing phylogenetic inference. In the age of genomes, single-gene trees, once used to test the predictions of endosymbiotic theory, now spawn new theories that stand to eventually replace endosymbiotic theory with descriptive, gene tree-based variants featuring supernumerary symbionts: prokaryotic partners distinct from the cornerstone trio and whose existence is inferred solely from single-gene trees. We reason that the endosymbiotic ancestors of mitochondria and chloroplasts brought into the eukaryotic—and plant and algal—lineage a genome-sized sample of genes from the proteobacterial and cyanobacterial pangenomes of their respective day and that, even if molecular phylogeny were artifact-free, sampling prokaryotic pangenomes through endosymbiotic gene transfer would lead to inherited chimerism. Recombination in prokaryotes (transduction, conjugation, transformation) differs from recombination in eukaryotes (sex). Prokaryotic recombination leads to pangenomes, and eukaryotic recombination leads to vertical inheritance. Viewed from the perspective of endosymbiotic theory, the critical transition at the eukaryote origin that allowed escape from Muller’s ratchet—the origin of eukaryotic recombination, or sex—might have required surprisingly little evolutionary innovation. PMID:25733873

  3. Wolbachia genome integrated in an insect chromosome: Evolution and fate of laterally transferred endosymbiont genes

    PubMed Central

    Nikoh, Naruo; Tanaka, Kohjiro; Shibata, Fukashi; Kondo, Natsuko; Hizume, Masahiro; Shimada, Masakazu; Fukatsu, Takema

    2008-01-01

    Recent accumulation of microbial genome data has demonstrated that lateral gene transfers constitute an important and universal evolutionary process in prokaryotes, while those in multicellular eukaryotes are still regarded as unusual, except for endosymbiotic gene transfers from mitochondria and plastids. Here we thoroughly investigated the bacterial genes derived from a Wolbachia endosymbiont on the nuclear genome of the beetle Callosobruchus chinensis. Exhaustive PCR detection and Southern blot analysis suggested that ∼30% of Wolbachia genes, in terms of the gene repertoire of wMel, are present on the insect nuclear genome. Fluorescent in situ hybridization located the transferred genes on the proximal region of the basal short arm of the X chromosome. Molecular evolutionary and other lines of evidence indicated that the transferred genes are probably derived from a single lateral transfer event. The transferred genes were, for the length examined, structurally disrupted, freed from functional constraints, and transcriptionally inactive. Hence, most, if not all, of the transferred genes have been pseudogenized. Notwithstanding this, the transferred genes were ubiquitously detected from Japanese and Taiwanese populations of C. chinensis, while the number of the transferred genes detected differed between the populations. The transferred genes were not detected from congenic beetle species, indicating that the transfer event occurred after speciation of C. chinensis, which was estimated to be one or several million years ago. These features of the laterally transferred endosymbiont genes are compared with the evolutionary patterns of mitochondrial and plastid genome fragments acquired by nuclear genomes through recent endosymbiotic gene transfers. PMID:18073380

  4. Innate Functions of Immunoglobulin M Lessen Liver Gene Transfer with Helper-Dependent Adenovirus

    PubMed Central

    Unzu, Carmen; Morales-Kastresana, Aizea; Sampedro, Ana; Serrano-Mendioroz, Irantzu; Azpilikueta, Arantza; Ochoa, María Carmen; Dubrot, Juan; Martínez-Ansó, Eduardo

    2014-01-01

    The immune system poses obstacles to viral vectors, even in the first administration to preimmunized hosts. We have observed that the livers of B cell-deficient mice were more effectively transduced by a helper-dependent adenovirus serotype-5 (HDA) vector than those of WT mice. This effect was T-cell independent as shown in athymic mice. Passive transfer of the serum from adenovirus-naïve WT to Rag1KO mice resulted in a reduction in gene transfer that was traced to IgM purified from serum of adenovirus-naïve mice. To ascribe the gene transfer inhibition activity to either adenoviral antigen-specific or antigen-unspecific functions of IgM, we used a monoclonal IgM antibody of unrelated specificity. Both the polyclonal and the irrelevant monoclonal IgM inhibited gene transfer by the HDA vector to either cultured hepatocellular carcinoma cells or to the liver of mice in vivo. Adsorption of polyclonal or monoclonal IgMs to viral capsids was revealed by ELISAs on adenovirus-coated plates. These observations indicate the existence of an inborn IgM mechanism deployed against a prevalent virus to reduce early post-infection viremia. In conclusion, innate IgM binding to adenovirus serotype-5 capsids restrains gene-transfer and offers a mechanism to be targeted for optimization of vector dosage in gene therapy with HDA vectors. PMID:24465560

  5. Innate functions of immunoglobulin M lessen liver gene transfer with helper-dependent adenovirus.

    PubMed

    Unzu, Carmen; Melero, Ignacio; Morales-Kastresana, Aizea; Sampedro, Ana; Serrano-Mendioroz, Irantzu; Azpilikueta, Arantza; Ochoa, María Carmen; Dubrot, Juan; Martínez-Ansó, Eduardo; Fontanellas, Antonio

    2014-01-01

    The immune system poses obstacles to viral vectors, even in the first administration to preimmunized hosts. We have observed that the livers of B cell-deficient mice were more effectively transduced by a helper-dependent adenovirus serotype-5 (HDA) vector than those of WT mice. This effect was T-cell independent as shown in athymic mice. Passive transfer of the serum from adenovirus-naïve WT to Rag1KO mice resulted in a reduction in gene transfer that was traced to IgM purified from serum of adenovirus-naïve mice. To ascribe the gene transfer inhibition activity to either adenoviral antigen-specific or antigen-unspecific functions of IgM, we used a monoclonal IgM antibody of unrelated specificity. Both the polyclonal and the irrelevant monoclonal IgM inhibited gene transfer by the HDA vector to either cultured hepatocellular carcinoma cells or to the liver of mice in vivo. Adsorption of polyclonal or monoclonal IgMs to viral capsids was revealed by ELISAs on adenovirus-coated plates. These observations indicate the existence of an inborn IgM mechanism deployed against a prevalent virus to reduce early post-infection viremia. In conclusion, innate IgM binding to adenovirus serotype-5 capsids restrains gene-transfer and offers a mechanism to be targeted for optimization of vector dosage in gene therapy with HDA vectors.

  6. Horizontal transfer of carbohydrate metabolism genes into ectomycorrhizal Amanita.

    PubMed

    Chaib De Mares, Maryam; Hess, Jaqueline; Floudas, Dimitrios; Lipzen, Anna; Choi, Cindy; Kennedy, Megan; Grigoriev, Igor V; Pringle, Anne

    2015-03-01

    The genus Amanita encompasses both symbiotic, ectomycorrhizal fungi and asymbiotic litter decomposers; all species are derived from asymbiotic ancestors. Symbiotic species are no longer able to degrade plant cell walls. The carbohydrate esterases family 1 (CE1s) is a diverse group of enzymes involved in carbon metabolism, including decomposition and carbon storage. CE1 genes of the ectomycorrhizal A. muscaria appear diverged from all other fungal homologues, and more similar to CE1s of bacteria, suggesting a horizontal gene transfer (HGT) event. In order to test whether AmanitaCE1s were acquired horizontally, we built a phylogeny of CE1s collected from across the tree of life, and describe the evolution of CE1 genes among Amanita and relevant lineages of bacteria. CE1s of symbiotic Amanita were very different from CE1s of asymbiotic Amanita, and are more similar to bacterial CE1s. The protein structure of one CE1 gene of A. muscaria matched a depolymerase that degrades the carbon storage molecule poly((R)-3-hydroxybutyrate) (PHB). Asymbiotic Amanita do not carry sequence or structural homologues of these genes. The CE1s acquired through HGT may enable novel metabolisms, or play roles in signaling or defense. This is the first evidence for the horizontal transfer of carbohydrate metabolism genes into ectomycorrhizal fungi.

  7. Human gene transfer: Characterization of human tumor-infiltrating lymphocytes as vehicles for retroviral-mediated gene transfer in man

    SciTech Connect

    Kasid, A.; Morecki, S.; Aebersold, P.; Cornetta, K.; Culver, K.; Freeman, S.; Director, E.; Lotze, M.T.; Blaese, R.M.; Anderson, W.F.; Rosenberg, S.A. )

    1990-01-01

    Tumor-infiltrating lymphocytes (TILs) are cells generated from tumor suspensions cultured in interleukin 2 that can mediate cancer regression when adoptively transferred into mice or humans. Since TILs proliferate rapidly in vitro, recirculate, and preferentially localize at the tumor site in vivo, they provide an attractive model for delivery of exogenous genetic material into man. To determine whether efficient gene transfer into TILs is feasible. The authors transduced human TILs with the bacterial gene for neomycin-resistance (Neo{sup R}) using the retroviral vector N2. The transduced TIL populations were stable and polyclonal with respect to the intact Neo{sup R} gene integration and expressed high levels of neomycin phosphotransferase activity. The Neo{sup R} gene insertion did not alter the in vitro growth pattern and interleukin 2 dependence of the transduced TILs. Analyses of T-cell receptor gene rearrangement for {beta}- and {gamma}-chain genes revealed the oligoclonal nature of the TIL populations with no major change in the DNA rearrangement patterns or the levels of mRNA expression of the {beta} and {gamma} chains following transduction and selection of TILs in the neomycin analog G418. Human TILs expressed mRNA for tumor necrosis factors ({alpha} and {beta}) and interleukin 2 receptor P55. This pattern of cytokine-mRNA expression was not significantly altered following the transduction of TILs. The studies demonstrate the feasibility of TILs as suitable cellular vehicles for the introduction of therapeutic genes into patients receiving autologous TILs.

  8. Transfer of tetracycline resistance genes with aggregation substance in food-borne Enterococcus faecalis.

    PubMed

    Choi, Jong-Mi; Woo, Gun-Jo

    2015-04-01

    Enterococcus faecalis has the ability to conjugate with the aid of aggregation substance (AS) and inducible sex pheromones to exchange genetic elements in food matrix. To evaluate the food safety condition and the transferable factor, 250 tetracycline-resistant food-borne E. faecalis were collected in Korea. Among the isolates, a majority of tetracycline-resistant isolates (49.6 %) harbored both the tet(M) and tet(L) genes together, followed by tet(M) (19.6 %), and tet(L) (6.8 %) alone. Also, we found the combination of tet(L)/tet(M)/tet(O) or tet(M)/tet(O). We identified two tet(S) genes including the isolate carrying tet(M) + tet(S) genes. Additionally, most E. faecalis were positive for cpd and ccf (both 96.8 %) followed by cob (57.2 %). Through mating experiments, we confirmed E. faecalis possessing the Int-Tn gene and/or any AS gene successfully transferred tet genes to JH2-2 E. faecalis, whereas neither E. faecalis carrying AS genes nor the Int-Tn gene showed the conjugation. Pulsed-field gel electrophoresis results supported a distinct pattern, implying transfer of genetic information. Our study revealed a high occurrence of tetracycline resistance genes in E. faecalis from various foods. The widespread dissemination of tetracycline resistance genes would be promoted to transfer tetracycline resistance genes by pheromone-mediated conjugation systems.

  9. Frequency of horizontal gene transfer of a large catabolic plasmid (pJP4) in soil.

    PubMed

    Neilson, J W; Josephson, K L; Pepper, I L; Arnold, R B; Di Giovanni, G D; Sinclair, N A

    1994-11-01

    Limited work has been done to assess the bioremediation potential of transfer of plasmid-borne degradative genes from introduced to indigenous organisms in the environment. Here we demonstrate the transfer by conjugation of the catabolic plasmid pJP4, using a model system with donor and recipient organisms. The donor organism was Alcaligenes eutrophus JMP134 and the recipient organism was Variovorax paradoxus isolated from a toxic waste site. Plasmid pJP4 contains genes for mercury resistance and 2,4-dichlorophenoxyacetic (2,4-D) acid degradation. A transfer frequency of approximately 1/10(3) donor and recipient cells (parent cells) was observed on solid agar media, decreasing to 1/10(5) parent cells in sterile soil and finally 1/10(6) parent cells in 2,4-D-amended, nonsterile soil. Presumptive transconjugants were confirmed to be resistant to Hg, to be capable of degrading 2,4-D, and to contain a plasmid of size comparable to that of pJP4. In addition, we confirmed the transfer through PCR amplifications of the tfdB gene. Although transfer of pJP4 did occur at a high frequency in pure culture, the rate was significantly decreased by the introduction of abiotic (sterile soil) and biotic (nonsterile soil) stresses. An evaluation of the data from this model system implies that the reliance on plasmid transfer from a donor organism as a remediative strategy has limited potential.

  10. Phosphatidylserine immobilization of lentivirus for localized gene transfer

    PubMed Central

    Shin, Seungjin; Tuinstra, Hannah M.; Salvay, David M.; Shea, Lonnie D.

    2010-01-01

    Localized and efficient gene transfer can be promoted by exploiting the interaction between the vector and biomaterial. Regulation of the vector-material interaction was investigated by capitalizing on the binding between lentivirus and phosphatidylserine (PS), a component of the plasma membrane. PS was incorporated into microspheres composed of the copolymers of lactide and glycolide (PLG) using an emulsion process. Increasing the weight ratio of PS to PLG led to a greater incorporation of PS. Lentivirus, but not adenovirus, associated with PS-PLG microspheres, and binding was specific to PS relative to PLG alone or PLG modified with phosphatidylcholine. Immobilized lentivirus produced large numbers of transduced cells, and increased transgene expression relative to virus alone. Microspheres were subsequently formed into porous tissue engineering scaffolds, with retention of lentivirus binding. Lentivirus immobilization resulted in long-term and localized expression within a subcutaneously implanted scaffold. Microspheres were also formed into multiple channel bridges for implantation into the spinal cord. Lentivirus delivery from the bridge produced maximal expression at the implant and a gradient of expression rostrally and caudally. This specific binding of lentiviral vectors to biomaterial scaffolds may provide a versatile tool for numerous applications in regenerative medicine or within model systems that investigate tissue development. PMID:20206382

  11. Design Criteria for Bagless Transfer System (BTS) Packaging System

    SciTech Connect

    RISENMAY, H.R.

    2000-04-26

    This document provides the criteria for the design and installation of a Bagless Transfer System (BTS); Blend, Sieve and Balance Equipment; and Supercritical Fluid Extraction System (SFE). The project consists of 3 major modules: (1) Bagless Transfer System (BTS) Module; (2) Blend, Sieve and Balance Equipment; and (3) Supercritical Fluid Extraction (SFE) Module.

  12. Transfer Function Control for Biometric Monitoring System

    NASA Technical Reports Server (NTRS)

    Chmiel, Alan J. (Inventor); Humphreys, Bradley T. (Inventor); Grodinsky, Carlos M. (Inventor)

    2015-01-01

    A modular apparatus for acquiring biometric data may include circuitry operative to receive an input signal indicative of a biometric condition, the circuitry being configured to process the input signal according to a transfer function thereof and to provide a corresponding processed input signal. A controller is configured to provide at least one control signal to the circuitry to programmatically modify the transfer function of the modular system to facilitate acquisition of the biometric data.

  13. Characterization of an Ancient Lepidopteran Lateral Gene Transfer

    PubMed Central

    Wheeler, David; Redding, Amanda J.; Werren, John H.

    2013-01-01

    Bacteria to eukaryote lateral gene transfers (LGT) are an important potential source of material for the evolution of novel genetic traits. The explosion in the number of newly sequenced genomes provides opportunities to identify and characterize examples of these lateral gene transfer events, and to assess their role in the evolution of new genes. In this paper, we describe an ancient lepidopteran LGT of a glycosyl hydrolase family 31 gene (GH31) from an Enterococcus bacteria. PCR amplification between the LGT and a flanking insect gene confirmed that the GH31 was integrated into the Bombyx mori genome and was not a result of an assembly error. Database searches in combination with degenerate PCR on a panel of 7 lepidopteran families confirmed that the GH31 LGT event occurred deep within the Order approximately 65–145 million years ago. The most basal species in which the LGT was found is Plutella xylostella (superfamily: Yponomeutoidea). Array data from Bombyx mori shows that GH31 is expressed, and low dN/dS ratios indicates the LGT coding sequence is under strong stabilizing selection. These findings provide further support for the proposition that bacterial LGTs are relatively common in insects and likely to be an underappreciated source of adaptive genetic material. PMID:23533610

  14. The electron transfer system of syntrophically grown Desulfovibrio vulgaris.

    PubMed

    Walker, Christopher B; He, Zhili; Yang, Zamin K; Ringbauer, Joseph A; He, Qiang; Zhou, Jizhong; Voordouw, Gerrit; Wall, Judy D; Arkin, Adam P; Hazen, Terry C; Stolyar, Sergey; Stahl, David A

    2009-09-01

    Interspecies hydrogen transfer between organisms producing and consuming hydrogen promotes the decomposition of organic matter in most anoxic environments. Although syntrophic coupling between hydrogen producers and consumers is a major feature of the carbon cycle, mechanisms for energy recovery at the extremely low free energies of reactions typical of these anaerobic communities have not been established. In this study, comparative transcriptional analysis of a model sulfate-reducing microbe, Desulfovibrio vulgaris Hildenborough, suggested the use of alternative electron transfer systems dependent on growth modality. During syntrophic growth on lactate with a hydrogenotrophic methanogen, numerous genes involved in electron transfer and energy generation were upregulated in D. vulgaris compared with their expression in sulfate-limited monocultures. In particular, genes coding for the putative membrane-bound Coo hydrogenase, two periplasmic hydrogenases (Hyd and Hyn), and the well-characterized high-molecular-weight cytochrome (Hmc) were among the most highly expressed and upregulated genes. Additionally, a predicted operon containing genes involved in lactate transport and oxidation exhibited upregulation, further suggesting an alternative pathway for electrons derived from lactate oxidation during syntrophic growth. Mutations in a subset of genes coding for Coo, Hmc, Hyd, and Hyn impaired or severely limited syntrophic growth but had little effect on growth via sulfate respiration. These results demonstrate that syntrophic growth and sulfate respiration use largely independent energy generation pathways and imply that to understand microbial processes that sustain nutrient cycling, lifestyles not captured in pure culture must be considered.

  15. The electron transfer system of synthrophically grown desulfovibrio vulgaris

    SciTech Connect

    Walker, Christopher; He, Zhili; Yang, Zamin Koo; Ringbauer, Joseph; HE, Qiang; Zhou, Jizhong; Voordouw, Gerrit; Wall, Judy; Arkin, Adam; Hazen, Terry; Stolyar, Sergey; Stahl, David

    2009-01-01

    Interspecies hydrogen transfer between organisms producing and consuming hydrogen promotes the decomposition of organic matter in most anoxic environments. Although syntrophic coupling between hydrogen producers and consumers is a major feature of the carbon cycle, mechanisms for energy recovery at the extremely low free energies of reactions typical of these anaerobic communities have not been established. In this study, comparative transcriptional analysis of a model sulfate-reducing microbe, Desulfovibrio vulgaris Hildenborough, suggested the use of alternative electron transfer systems dependent on growth modality. During syntrophic growth on lactate with a hydrogenotrophic methanogen, numerous genes involved in electron transfer and energy generation were upregulated in D. vulgaris compared with their expression in sulfate-limited monocultures. In particular, genes coding for the putative membrane-bound Coo hydrogenase, two periplasmic hydrogenases (Hyd and Hyn), and the well-characterized high-molecular-weight cytochrome (Hmc) were among the most highly expressed and upregulated genes. Additionally, a predicted operon containing genes involved in lactate transport and oxidation exhibited upregulation, further suggesting an alternative pathway for electrons derived from lactate oxidation during syntrophic growth. Mutations in a subset of genes coding for Coo, Hmc, Hyd, and Hyn impaired or severely limited syntrophic growth but had little effect on growth via sulfate respiration. These results demonstrate that syntrophic growth and sulfate respiration use largely independent energy generation pathways and imply that to understand microbial processes that sustain nutrient cycling, lifestyles not captured in pure culture must be considered.

  16. Gene transfer from a parasitic flowering plant to a fern.

    PubMed

    Davis, Charles C; Anderson, William R; Wurdack, Kenneth J

    2005-11-07

    The rattlesnake fern (Botrychium virginianum (L.) Sw.) is obligately mycotrophic and widely distributed across the northern hemisphere. Three mitochondrial gene regions place this species with other ferns in Ophioglossaceae, while two regions place it as a member of the largely parasitic angiosperm order Santalales (sandalwoods and mistletoes). These discordant phylogenetic placements suggest that part of the genome in B. virginianum was acquired by horizontal gene transfer (HGT), perhaps from root-parasitic Loranthaceae. These transgenes are restricted to B. virginianum and occur across the range of the species. Molecular and life-history traits indicate that the transfer preceded the global expansion of B. virginianum, and that the latter may have happened very rapidly. This is the first report of HGT from an angiosperm to a fern, through either direct parasitism or the mediation of interconnecting fungal symbionts.

  17. Gene transfer from a parasitic flowering plant to a fern

    PubMed Central

    Davis, Charles C; Anderson, William R; Wurdack, Kenneth J

    2005-01-01

    The rattlesnake fern (Botrychium virginianum (L.) Sw.) is obligately mycotrophic and widely distributed across the northern hemisphere. Three mitochondrial gene regions place this species with other ferns in Ophioglossaceae, while two regions place it as a member of the largely parasitic angiosperm order Santalales (sandalwoods and mistletoes). These discordant phylogenetic placements suggest that part of the genome in B. virginianum was acquired by horizontal gene transfer (HGT), perhaps from root-parasitic Loranthaceae. These transgenes are restricted to B. virginianum and occur across the range of the species. Molecular and life-history traits indicate that the transfer preceded the global expansion of B. virginianum, and that the latter may have happened very rapidly. This is the first report of HGT from an angiosperm to a fern, through either direct parasitism or the mediation of interconnecting fungal symbionts. PMID:16191635

  18. Maximizing energy transfer in vibrofluidized granular systems.

    PubMed

    Windows-Yule, C R K; Rosato, A D; Parker, D J; Thornton, A R

    2015-05-01

    Using discrete particle simulations validated by experimental data acquired using the positron emission particle tracking technique, we study the efficiency of energy transfer from a vibrating wall to a system of discrete, macroscopic particles. We demonstrate that even for a fixed input energy from the wall, energy conveyed to the granular system under excitation may vary significantly dependent on the frequency and amplitude of the driving oscillations. We investigate the manner in which the efficiency with which energy is transferred to the system depends on the system variables and determine the key control parameters governing the optimization of this energy transfer. A mechanism capable of explaining our results is proposed, and the implications of our findings in the research field of granular dynamics as well as their possible utilization in industrial applications are discussed.

  19. [Advances in molecular mechanisms of bacterial resistance caused by stress-induced transfer of resistance genes--a review].

    PubMed

    Sun, Dongchang; Wang, Bing; Zhu, Lihong

    2013-07-04

    The transfer of resistance gene is one of the most important causes of bacterial resistance. Recent studies reveal that stresses induce the transfer of antibiotic resistance gene through multiple mechanisms. DNA damage stresses trigger bacterial SOS response and induce the transfer of resistance gene mediated by conjugative DNA. Antibiotic stresses induce natural bacterial competence for transformation in some bacteria which lack the SOS system. In addition, our latest studies show that the general stress response regulator RpoS regulates a novel type of resistance gene transfer which is mediated by double-stranded plasmid DNA and occurs exclusively on the solid surface. In this review, we summarized recent advances in SOS dependent and independent stress-induced DNA transfer which is mediated by conjugation and transformation respectively, and the transfer of double-stranded plasmid DNA on the solid surface which is regulated by RpoS. We propose that future work should address how stresses activate the key regulators and how these regulators control the expression of gene transfer related genes. Answers to the above questions would pave the way for searching for candidate targets for controlling bacterial resistance resulted from the transfer of antibiotic genes.

  20. Risks from GMOs due to horizontal gene transfer.

    PubMed

    Keese, Paul

    2008-01-01

    Horizontal gene transfer (HGT) is the stable transfer of genetic material from one organism to another without reproduction or human intervention. Transfer occurs by the passage of donor genetic material across cellular boundaries, followed by heritable incorporation to the genome of the recipient organism. In addition to conjugation, transformation and transduction, other diverse mechanisms of DNA and RNA uptake occur in nature. The genome of almost every organism reveals the footprint of many ancient HGT events. Most commonly, HGT involves the transmission of genes on viruses or mobile genetic elements. HGT first became an issue of public concern in the 1970s through the natural spread of antibiotic resistance genes amongst pathogenic bacteria, and more recently with commercial production of genetically modified (GM) crops. However, the frequency of HGT from plants to other eukaryotes or prokaryotes is extremely low. The frequency of HGT to viruses is potentially greater, but is restricted by stringent selection pressures. In most cases the occurrence of HGT from GM crops to other organisms is expected to be lower than background rates. Therefore, HGT from GM plants poses negligible risks to human health or the environment.

  1. Horizontal Gene Transfer is a Significant Driver of Gene Innovation in Dinoflagellates

    PubMed Central

    Wisecaver, Jennifer H.; Brosnahan, Michael L.; Hackett, Jeremiah D.

    2013-01-01

    The dinoflagellates are an evolutionarily and ecologically important group of microbial eukaryotes. Previous work suggests that horizontal gene transfer (HGT) is an important source of gene innovation in these organisms. However, dinoflagellate genomes are notoriously large and complex, making genomic investigation of this phenomenon impractical with currently available sequencing technology. Fortunately, de novo transcriptome sequencing and assembly provides an alternative approach for investigating HGT. We sequenced the transcriptome of the dinoflagellate Alexandrium tamarense Group IV to investigate how HGT has contributed to gene innovation in this group. Our comprehensive A. tamarense Group IV gene set was compared with those of 16 other eukaryotic genomes. Ancestral gene content reconstruction of ortholog groups shows that A. tamarense Group IV has the largest number of gene families gained (314–1,563 depending on inference method) relative to all other organisms in the analysis (0–782). Phylogenomic analysis indicates that genes horizontally acquired from bacteria are a significant proportion of this gene influx, as are genes transferred from other eukaryotes either through HGT or endosymbiosis. The dinoflagellates also display curious cases of gene loss associated with mitochondrial metabolism including the entire Complex I of oxidative phosphorylation. Some of these missing genes have been functionally replaced by bacterial and eukaryotic xenologs. The transcriptome of A. tamarense Group IV lends strong support to a growing body of evidence that dinoflagellate genomes are extraordinarily impacted by HGT. PMID:24259313

  2. Efficient transfer of two large secondary metabolite pathway gene clusters into heterologous hosts by transposition

    PubMed Central

    Fu, Jun; Wenzel, Silke C.; Perlova, Olena; Wang, Junping; Gross, Frank; Tang, Zhiru; Yin, Yulong; Stewart, A. Francis; Zhang, Youming

    2008-01-01

    Horizontal gene transfer by transposition has been widely used for transgenesis in prokaryotes. However, conjugation has been preferred for transfer of large transgenes, despite greater restrictions of host range. We examine the possibility that transposons can be used to deliver large transgenes to heterologous hosts. This possibility is particularly relevant to the expression of large secondary metabolite gene clusters in various heterologous hosts. Recently, we showed that the engineering of large gene clusters like type I polyketide/nonribosomal peptide pathways for heterologous expression is no longer a bottleneck. Here, we apply recombineering to engineer either the epothilone (epo) or myxochromide S (mchS) gene cluster for transpositional delivery and expression in heterologous hosts. The 58-kb epo gene cluster was fully reconstituted from two clones by stitching. Then, the epo promoter was exchanged for a promoter active in the heterologous host, followed by engineering into the MycoMar transposon. A similar process was applied to the mchS gene cluster. The engineered gene clusters were transferred and expressed in the heterologous hosts Myxococcus xanthus and Pseudomonas putida. We achieved the largest transposition yet reported for any system and suggest that delivery by transposon will become the method of choice for delivery of large transgenes, particularly not only for metabolic engineering but also for general transgenesis in prokaryotes and eukaryotes. PMID:18701643

  3. Phylogeographic support for horizontal gene transfer involving sympatric bruchid species

    PubMed Central

    Alvarez, Nadir; Benrey, Betty; Hossaert-McKey, Martine; Grill, Andrea; McKey, Doyle; Galtier, Nicolas

    2006-01-01

    Background We report on the probable horizontal transfer of a mitochondrial gene, cytb, between species of Neotropical bruchid beetles, in a zone where these species are sympatric. The bruchid beetles Acanthoscelides obtectus, A. obvelatus, A. argillaceus and Zabrotes subfasciatus develop on various bean species in Mexico. Whereas A. obtectus and A. obvelatus develop on Phaseolus vulgaris in the Mexican Altiplano, A. argillaceus feeds on P. lunatus in the Pacific coast. The generalist Z. subfasciatus feeds on both bean species, and is sympatric with A. obtectus and A. obvelatus in the Mexican Altiplano, and with A. argillaceus in the Pacific coast. In order to assess the phylogenetic position of these four species, we amplified and sequenced one nuclear (28S rRNA) and two mitochondrial (cytb, COI) genes. Results Whereas species were well segregated in topologies obtained for COI and 28S rRNA, an unexpected pattern was obtained in the cytb phylogenetic tree. In this tree, individuals from A. obtectus and A. obvelatus, as well as Z. subfasciatus individuals from the Mexican Altiplano, clustered together in a unique little variable monophyletic unit. In contrast, A. argillaceus and Z. subfasciatus individuals from the Pacific coast clustered in two separated clades, identically to the pattern obtained for COI and 28S rRNA. An additional analysis showed that Z. subfasciatus individuals from the Mexican Altiplano also possessed the cytb gene present in individuals of this species from the Pacific coast. Zabrotes subfasciatus individuals from the Mexican Altiplano thus demonstrated two cytb genes, an "original" one and an "infectious" one, showing 25% of nucleotide divergence. The "infectious" cytb gene seems to be under purifying selection and to be expressed in mitochondria. Conclusion The high degree of incongruence of the cytb tree with patterns for other genes is discussed in the light of three hypotheses: experimental contamination, hybridization, and

  4. [Polymeric nanoparticles with therapeutic gene for gene therapy: I. Preparation and in vivo gene transfer study].

    PubMed

    Yang, Jing; Song, Cunxian; Sun, Hongfan; Wu, Li; Tang, Lina; Leng, Xigang; Wang, Pengyan; Xu, Yiyao; Li, Yongjun; Guan, Heng

    2005-06-01

    VEGF nanoparticle (VEGF-NP) was prepared by a multi-emulsification technique using a biodegradable poly-dl-lactic-co-glycolic (PLGA) as matrix material. The nanoparticles were characterized for size, VEGF loading capacity, and in vitro release. VEGF-NP and naked VEGF plasmid were intramuscularly injected into the ischemia site of the rabbit chronic hindlimb ischemia model and the efficiency of VEGF-NP as gene delivery carrier for gene therapy in animal model was evaluated. Gene therapuetic effect was assessed evaluated by RT-PCR, immunohistochemistry and angiography assay. The average size of VEGF-NP was around 300 nm. The encapsulation efficiency of VEGF was above 96%. Loading amount of VEGF in the nanoparticles was about 4%. In vitro, nanoparticles maintained sustained-release of VEGF for two weeks. Two weeks post gene injection the capillary density in VEGF-NP group (81.22 per mm2) was significantly higher than that in control group (29.54 mm2). RT-PCR results showed greatly higher VEGF expression in VEGF-NP group (31.79au * mm) than that in naked VEGF group (9.15 au * mm). As a carrier system for gene therapy in animal model, VEGF-NP is much better than naked DNA plasmid. The results demonstrate great possibility of using NP carrier in human gene therapy.

  5. Energy transfer in the solar system

    NASA Astrophysics Data System (ADS)

    Jelbring, H.

    2013-12-01

    Different types of energy transfer are presented from the literature and are approached and commented on. It follows from these articles that energy transfer in addition to solar irradiation is less well understood by contemporary scientist. The transformation of energy between kinetic and potential energy in planetary orbits might be of crucial importance for understanding energy transfer between celestial bodies and the development of commensurabilities. There is evidence pointing to interactions (friction) between space and satellites producing volcanism. The reversible transfer of energy between the orbit of Moon and Earth's rotational energy is crucial to the creation of the 13.6-day and 27.3-day periods in both solar variables and Earth bound climate variables. It is hypothesized that the Earth-Moon system is modulating the sunspot numbers and creating both these periods, and that the great planets are responsible for the 11 yr solar cycle.

  6. Gene Transfer by Guanidinium-Cholesterol Cationic Lipids into Airway Epithelial Cells in vitro and in vivo

    NASA Astrophysics Data System (ADS)

    Oudrhiri, Noufissa; Vigneron, Jean-Pierre; Peuchmaur, Michel; Leclerc, Tony; Lehn, Jean-Marie; Lehn, Pierre

    1997-03-01

    Synthetic vectors represent an attractive alternative approach to viral vectors for gene transfer, in particular into airway epithelial cells for lung-directed gene therapy for cystic fibrosis. Having recently found that guanidinium-cholesterol cationic lipids are efficient reagents for gene transfer into mammalian cell lines in vitro, we have investigated their use for gene delivery into primary airway epithelial cells in vitro and in vivo. The results obtained indicate that the lipid bis (guanidinium)-tren-cholesterol (BGTC) can be used to transfer a reporter gene into primary human airway epithelial cells in culture. Furthermore, liposomes composed of BGTC and dioleoyl phosphatidylethanolamine (DOPE) are efficient for gene delivery to the mouse airway epithelium in vivo. Transfected cells were detected both in the surface epithelium and in submucosal glands. In addition, the transfection efficiency of BGTC/DOPE liposomes in vivo was quantitatively assessed by using the luciferase reporter gene system.

  7. Gene transfer into Solanum tuberosum via Rhizobium spp.

    PubMed

    Wendt, Toni; Doohan, Fiona; Winckelmann, Dominik; Mullins, Ewen

    2011-04-01

    Agrobacterium tumefaciens-mediated transformation (ATMT) is the preferred technique for gene transfer into crops. A major disadvantage of the technology remains the complexity of the patent landscape that surrounds ATMT which restricts its use for commercial applications. An alternative system has been described (Broothaerts et al. in Nature 433:629-633, 2005) detailing the propensity of three rhizobia to transform the model crop Arabidopsis thaliana, the non-food crop Nicotiana tabacum and, at a very low frequency, the monocotyledonous crop Oryza sativa. In this report we describe for the first time the genetic transformation of Solanum tuberosum using the non-Agrobacterium species Sinorhizobium meliloti, Rhizobium sp. NGR234 and Mesorhizobium loti. This was achieved by combining an optimal bacterium and host co-cultivation period with a low antibiotic regime during the callus and shoot induction stages. Using this optimized protocol the transformation frequency (calculated as % of shoots equipped with root systems with the ability to grow in rooting media supplemented with 25 μg/ml hygromycin) of the rhizobia strains was calculated at 4.72, 5.85 and 1.86% for S. meliloti, R. sp. NGR234 and M. loti respectively, compared to 47.6% for the A. tumefaciens control. Stable transgene integration and expression was confirmed via southern hybridisation, quantitative PCR analysis and histochemical screening of both leaf and/or tuber tissue. In light of the rapid advances in potato genomics, combined with the sequencing of the potato genome, the ability of alternative bacteria species to genetically transform this major food crop will provide a novel resource to the Solanaceae community as it continues to develop potato as both a food and non-food crop.

  8. Horizontal Gene Transfer Contributes to the Evolution of Arthropod Herbivory.

    PubMed

    Wybouw, Nicky; Pauchet, Yannick; Heckel, David G; Van Leeuwen, Thomas

    2016-06-27

    Within animals, evolutionary transition toward herbivory is severely limited by the hostile characteristics of plants. Arthropods have nonetheless counteracted many nutritional and defensive barriers imposed by plants and are currently considered as the most successful animal herbivores in terrestrial ecosystems. We gather a body of evidence showing that genomes of various plant feeding insects and mites possess genes whose presence can only be explained by horizontal gene transfer (HGT). HGT is the asexual transmission of genetic information between reproductively isolated species. Although HGT is known to have great adaptive significance in prokaryotes, its impact on eukaryotic evolution remains obscure. Here, we show that laterally transferred genes into arthropods underpin many adaptations to phytophagy, including efficient assimilation and detoxification of plant produced metabolites. Horizontally acquired genes and the traits they encode often functionally diversify within arthropod recipients, enabling the colonization of more host plant species and organs. We demonstrate that HGT can drive metazoan evolution by uncovering its prominent role in the adaptations of arthropods to exploit plants.

  9. Horizontal Gene Transfer Contributes to the Evolution of Arthropod Herbivory

    PubMed Central

    Wybouw, Nicky; Pauchet, Yannick; Heckel, David G.; Van Leeuwen, Thomas

    2016-01-01

    Within animals, evolutionary transition toward herbivory is severely limited by the hostile characteristics of plants. Arthropods have nonetheless counteracted many nutritional and defensive barriers imposed by plants and are currently considered as the most successful animal herbivores in terrestrial ecosystems. We gather a body of evidence showing that genomes of various plant feeding insects and mites possess genes whose presence can only be explained by horizontal gene transfer (HGT). HGT is the asexual transmission of genetic information between reproductively isolated species. Although HGT is known to have great adaptive significance in prokaryotes, its impact on eukaryotic evolution remains obscure. Here, we show that laterally transferred genes into arthropods underpin many adaptations to phytophagy, including efficient assimilation and detoxification of plant produced metabolites. Horizontally acquired genes and the traits they encode often functionally diversify within arthropod recipients, enabling the colonization of more host plant species and organs. We demonstrate that HGT can drive metazoan evolution by uncovering its prominent role in the adaptations of arthropods to exploit plants. PMID:27307274

  10. Gene Therapy Inhibiting Neointimal Vascular Lesion: In vivo Transfer of Endothelial Cell Nitric Oxide Synthase Gene

    NASA Astrophysics Data System (ADS)

    von der Leyen, Heiko E.; Gibbons, Gary H.; Morishita, Ryuichi; Lewis, Neil P.; Zhang, Lunan; Nakajima, Masatoshi; Kaneda, Yasufumi; Cooke, John P.; Dzau, Victor J.

    1995-02-01

    It is postulated that vascular disease involves a disturbance in the homeostatic balance of factors regulating vascular tone and structure. Recent developments in gene transfer techniques have emerged as an exciting therapeutic option to treat vascular disease. Several studies have established the feasibility of direct in vivo gene transfer into the vasculature by using reporter genes such as β-galactosidase or luciferase. To date no study has documented therapeutic effects with in vivo gene transfer of a cDNA encoding a functional enzyme. This study tests the hypothesis that endothelium-derived nitric oxide is an endogenous inhibitor of vascular lesion formation. After denudation by balloon injury of the endothelium of rat carotid arteries, we restored endothelial cell nitric oxide synthase (ec-NOS) expression in the vessel wall by using the highly efficient Sendai virus/liposome in vivo gene transfer technique. ec-NOS gene transfection not only restored NO production to levels seen in normal untreated vessels but also increased vascular reactivity of the injured vessel. Neointima formation at day 14 after balloon injury was inhibited by 70%. These findings provide direct evidence that NO is an endogenous inhibitor of vascular lesion formation in vivo (by inhibiting smooth muscle cell proliferation and migration) and suggest the possibility of ec-NOS transfection as a potential therapeutic approach to treat neointimal hyperplasia.

  11. HGTree: database of horizontally transferred genes determined by tree reconciliation.

    PubMed

    Jeong, Hyeonsoo; Sung, Samsun; Kwon, Taehyung; Seo, Minseok; Caetano-Anollés, Kelsey; Choi, Sang Ho; Cho, Seoae; Nasir, Arshan; Kim, Heebal

    2016-01-04

    The HGTree database provides putative genome-wide horizontal gene transfer (HGT) information for 2472 completely sequenced prokaryotic genomes. This task is accomplished by reconstructing approximate maximum likelihood phylogenetic trees for each orthologous gene and corresponding 16S rRNA reference species sets and then reconciling the two trees under parsimony framework. The tree reconciliation method is generally considered to be a reliable way to detect HGT events but its practical use has remained limited because the method is computationally intensive and conceptually challenging. In this regard, HGTree (http://hgtree.snu.ac.kr) represents a useful addition to the biological community and enables quick and easy retrieval of information for HGT-acquired genes to better understand microbial taxonomy and evolution. The database is freely available and can be easily scaled and updated to keep pace with the rapid rise in genomic information.

  12. HGTree: database of horizontally transferred genes determined by tree reconciliation

    PubMed Central

    Jeong, Hyeonsoo; Sung, Samsun; Kwon, Taehyung; Seo, Minseok; Caetano-Anollés, Kelsey; Choi, Sang Ho; Cho, Seoae; Nasir, Arshan; Kim, Heebal

    2016-01-01

    The HGTree database provides putative genome-wide horizontal gene transfer (HGT) information for 2472 completely sequenced prokaryotic genomes. This task is accomplished by reconstructing approximate maximum likelihood phylogenetic trees for each orthologous gene and corresponding 16S rRNA reference species sets and then reconciling the two trees under parsimony framework. The tree reconciliation method is generally considered to be a reliable way to detect HGT events but its practical use has remained limited because the method is computationally intensive and conceptually challenging. In this regard, HGTree (http://hgtree.snu.ac.kr) represents a useful addition to the biological community and enables quick and easy retrieval of information for HGT-acquired genes to better understand microbial taxonomy and evolution. The database is freely available and can be easily scaled and updated to keep pace with the rapid rise in genomic information. PMID:26578597

  13. History of Polymeric Gene Delivery Systems.

    PubMed

    Zhang, Peng; Wagner, Ernst

    2017-04-01

    As an option for genetic disease treatment and an alternative for traditional cancer chemotherapy, gene therapy achieves significant attention. Nucleic acid delivery, however, remains a main challenge in human gene therapy. Polymer-based delivery systems offer a safer and promising route for therapeutic gene delivery. Over the past five decades, various cationic polymers have been optimized for increasingly effective nucleic acid transfer. This resulted in a chemical evolution of cationic polymers from the first-generation polycations towards bioinspired multifunctional sequence-defined polymers and nanocomposites. With the increasing of knowledge in molecular biological processes and rapid progress of macromolecular chemistry, further improvement of polymeric nucleic acid delivery systems will provide effective tool for gene-based therapy in the near future.

  14. Bacterial gene transfer by natural genetic transformation in the environment.

    PubMed Central

    Lorenz, M G; Wackernagel, W

    1994-01-01

    Natural genetic transformation is the active uptake of free DNA by bacterial cells and the heritable incorporation of its genetic information. Since the famous discovery of transformation in Streptococcus pneumoniae by Griffith in 1928 and the demonstration of DNA as the transforming principle by Avery and coworkers in 1944, cellular processes involved in transformation have been studied extensively by in vitro experimentation with a few transformable species. Only more recently has it been considered that transformation may be a powerful mechanism of horizontal gene transfer in natural bacterial populations. In this review the current understanding of the biology of transformation is summarized to provide the platform on which aspects of bacterial transformation in water, soil, and sediments and the habitat of pathogens are discussed. Direct and indirect evidence for gene transfer routes by transformation within species and between different species will be presented, along with data suggesting that plasmids as well as chromosomal DNA are subject to genetic exchange via transformation. Experiments exploring the prerequisites for transformation in the environment, including the production and persistence of free DNA and factors important for the uptake of DNA by cells, will be compiled, as well as possible natural barriers to transformation. The efficiency of gene transfer by transformation in bacterial habitats is possibly genetically adjusted to submaximal levels. The fact that natural transformation has been detected among bacteria from all trophic and taxonomic groups including archaebacteria suggests that transformability evolved early in phylogeny. Probable functions of DNA uptake other than gene acquisition will be discussed. The body of information presently available suggests that transformation has a great impact on bacterial population dynamics as well as on bacterial evolution and speciation. PMID:7968924

  15. The Electron Transfer System of Syntrophically Grown Desulfovibrio vulgaris

    SciTech Connect

    PBD; ENIGMA; GTL; VIMSS; Walker, Christopher B.; He, Zhili; Yang, Zamin K.; Ringbauer Jr., Joseph A.; He, Qiang; Zhou, Jizhong; Voordouw, Gerrit; Wall, Judy D.; Arkin, Adam P.; Hazen, Terry C.; Stolyar, Sergey; Stahl, David A.

    2009-06-22

    Interspecies hydrogen transfer between organisms producing and consuming hydrogen promotes the decomposition of organic matter in most anoxic environments. Although syntrophic couplings between hydrogen producers and consumers are a major feature of the carbon cycle, mechanisms for energy recovery at the extremely low free energies of reactions typical of these anaerobic communities have not been established. In this study, comparative transcriptional analysis of a model sulfate-reducing microbe, Desulfovibrio vulgaris Hildenborough, suggested the use of alternative electron transfer systems dependent upon growth modality. During syntrophic growth on lactate with a hydrogenotrophic methanogen, D. vulgaris up-regulated numerous genes involved in electron transfer and energy generation when compared with sulfate-limited monocultures. In particular, genes coding for the putative membrane-bound Coo hydrogenase, two periplasmic hydrogenases (Hyd and Hyn) and the well-characterized high-molecular weight cytochrome (Hmc) were among the most highly expressed and up-regulated. Additionally, a predicted operon coding for genes involved in lactate transport and oxidation exhibited up-regulation, further suggesting an alternative pathway for electrons derived from lactate oxidation during syntrophic growth. Mutations in a subset of genes coding for Coo, Hmc, Hyd and Hyn impaired or severely limited syntrophic growth but had little affect on growth via sulfate-respiration. These results demonstrate that syntrophic growth and sulfate-respiration use largely independent energy generation pathways and imply that understanding of microbial processes sustaining nutrient cycling must consider lifestyles not captured in pure culture.

  16. The electron transfer system of syntrophically grown Desulfovibrio vulgaris

    SciTech Connect

    Walker, C.B.; He, Z.; Yang, Z.K.; Ringbauer, Jr., J.A.; He, Q.; Zhou, J.; Voordouw, G.; Wall, J.D.; Arkin, A.P.; Hazen, T.C.; Stolyar, S.; Stahl, D.A.

    2009-05-01

    Interspecies hydrogen transfer between organisms producing and consuming hydrogen promotes the decomposition of organic matter in most anoxic environments. Although syntrophic couplings between hydrogen producers and consumers are a major feature of the carbon cycle, mechanisms for energy recovery at the extremely low free energies of reactions typical of these anaerobic communities have not been established. In this study, comparative transcriptional analysis of a model sulfate-reducing microbe, Desulfovibrio vulgaris Hildenborough, suggested the use of alternative electron transfer systems dependent upon growth modality. During syntrophic growth on lactate with a hydrogenotrophic methanogen, D. vulgaris up-regulated numerous genes involved in electron transfer and energy generation when compared with sulfate-limited monocultures. In particular, genes coding for the putative membrane-bound Coo hydrogenase, two periplasmic hydrogenases (Hyd and Hyn) and the well-characterized high-molecular weight cytochrome (Hmc) were among the most highly expressed and up-regulated. Additionally, a predicted operon coding for genes involved in lactate transport and oxidation exhibited up-regulation, further suggesting an alternative pathway for electrons derived from lactate oxidation during syntrophic growth. Mutations in a subset of genes coding for Coo, Hmc, Hyd and Hyn impaired or severely limited syntrophic growth but had little affect on growth via sulfate-respiration. These results demonstrate that syntrophic growth and sulfate-respiration use largely independent energy generation pathways and imply that understanding of microbial processes sustaining nutrient cycling must consider lifestyles not captured in pure culture.

  17. Plasmid transfer systems in the rhizobia.

    PubMed

    Ding, Hao; Hynes, Michael F

    2009-08-01

    Rhizobia are agriculturally important bacteria that can form nitrogen-fixing nodules on the roots of leguminous plants. Agricultural application of rhizobial inoculants can play an important role in increasing leguminous crop yields. In temperate rhizobia, genes involved in nodulation and nitrogen fixation are usually located on one or more large plasmids (pSyms) or on symbiotic islands. In addition, other large plasmids of rhizobia carry genes that are beneficial for survival and competition of rhizobia in the rhizosphere. Conjugative transfer of these large plasmids thus plays an important role in the evolution of rhizobia. Therefore, understanding the mechanism of conjugative transfer of large rhizobial plasmids provides foundations for maintaining, monitoring, and predicting the behaviour of these plasmids during field release events. In this minireview, we summarize two types of known rhizobial conjugative plasmids, including quorum sensing regulated plasmids and RctA-repressed plasmids. We provide evidence for the existence of a third type of conjugative plasmid, including pRleVF39c in Rhizobium leguminosarum bv. viciae strain VF39SM, and we provide a comparison of the different types of conjugation genes found in members of the rhizobia that have had their genomes sequenced so far.

  18. Orbital transfer systems for lunar missions

    NASA Astrophysics Data System (ADS)

    Koelle, Dietrich E.; Obersteiner, Michael

    For future Lunar mission operations basically two types of transfer systems can be conceived (a) the conventional direct-injection expendable vehicle approach, and (b) the re-usable orbit-to-orbit (LEO-LLO-LEO round-trip system. The second solution has been studied in Europe in the SPACE TUG Phase A Study in 1971/72 for NASA as a fully reusable system and this is compared to a more recent NASA-MSFC transfer vehicle concept with expendable propellant tanks and aerobraking. The results of an analysis on mission opportunities for LEO-LLO transfers is presented. A European option is also shown for mode (a), using the ARIANE 5 for lunar missions with an optimized cryogenic transfer vehicle, derived from the H.10 Stage (ARIANE 4) plus a lunar vehicle derived from the MMII-CRAF propulsion module using the 27.5 kN engine from the ARIANE L.9 Stage which could land a 3.4 ton vehicle on the lunar surface. Finally a completely new option (c) is discussed: the use of a ballistic SSTO vehicle for lunar landing missions after refuelling in LEO. This seems to be a highly effective single transportation element approach compared to the seven elements SATURN V-APOLLO-LM system of the 60ies.

  19. Information transfer in the National Airspace System

    NASA Technical Reports Server (NTRS)

    Lee, Alfred T.

    1988-01-01

    An informal overview is given of the work in progress and the planned work in the area of information transfer that specifically addresses human factors issues in National Airspace System (NAS). The issues of how weather information will be displayed on the flight deck, the development of appropriate decision making technology, and digital datalink transmission are also briefly discussed.

  20. Promoting Transfer by Grounding Complex Systems Principles

    ERIC Educational Resources Information Center

    Goldstone, Robert L.; Wilensky, Uri

    2008-01-01

    Understanding scientific phenomena in terms of complex systems principles is both scientifically and pedagogically important. Situations from different disciplines of science are often governed by the same principle, and so promoting knowledge transfer across disciplines makes valuable cross-fertilization and scientific unification possible.…

  1. System Transfer, Education, and Development in Mozambique

    ERIC Educational Resources Information Center

    Cossa, Jose

    2011-01-01

    In this study the author used conceptual historical method to assess the phenomenon of system transfer and the association between education and development in Mozambique. The assessment was administered through critical analysis of documents pertaining to the Salazar (1924-1966), Machel (1975-1986), and Chissano (1986-2005) administrations. The…

  2. Adaptive Horizontal Gene Transfers between Multiple Cheese-Associated Fungi.

    PubMed

    Ropars, Jeanne; Rodríguez de la Vega, Ricardo C; López-Villavicencio, Manuela; Gouzy, Jérôme; Sallet, Erika; Dumas, Émilie; Lacoste, Sandrine; Debuchy, Robert; Dupont, Joëlle; Branca, Antoine; Giraud, Tatiana

    2015-10-05

    Domestication is an excellent model for studies of adaptation because it involves recent and strong selection on a few, identified traits [1-5]. Few studies have focused on the domestication of fungi, with notable exceptions [6-11], despite their importance to bioindustry [12] and to a general understanding of adaptation in eukaryotes [5]. Penicillium fungi are ubiquitous molds among which two distantly related species have been independently selected for cheese making-P. roqueforti for blue cheeses like Roquefort and P. camemberti for soft cheeses like Camembert. The selected traits include morphology, aromatic profile, lipolytic and proteolytic activities, and ability to grow at low temperatures, in a matrix containing bacterial and fungal competitors [13-15]. By comparing the genomes of ten Penicillium species, we show that adaptation to cheese was associated with multiple recent horizontal transfers of large genomic regions carrying crucial metabolic genes. We identified seven horizontally transferred regions (HTRs) spanning more than 10 kb each, flanked by specific transposable elements, and displaying nearly 100% identity between distant Penicillium species. Two HTRs carried genes with functions involved in the utilization of cheese nutrients or competition and were found nearly identical in multiple strains and species of cheese-associated Penicillium fungi, indicating recent selective sweeps; they were experimentally associated with faster growth and greater competitiveness on cheese and contained genes highly expressed in the early stage of cheese maturation. These findings have industrial and food safety implications and improve our understanding of the processes of adaptation to rapid environmental changes.

  3. Adenovirus dodecahedron, a new vector for human gene transfer.

    PubMed

    Fender, P; Ruigrok, R W; Gout, E; Buffet, S; Chroboczek, J

    1997-01-01

    Recombinant adenovirus is one of most efficient delivery vehicles for gene therapy. However, the initial enthusiasm for the use of recombinant adenovirus for gene therapy has been tempered by strong immune responses that develop to the virus and virus-infected cells. Even though recombinant adenoviruses are replication-defective, they introduce into the recipient cell, together with the gene of interest, viral genetes that might lead to fortuitous recombination if the recipient is infected by wild-type adenovirus. We propose the use of a dodecahedron made of adenovirus pentons or penton bases as an alternative vector for human gene therapy. The penton is a complex of two oligomeric proteins, a penton base and fiber, involved in the cell attachment, internalization, and liberation of virus into the cytoplasm. The dodecahedron retains many of the advantages of adenovirus for gene transfer such as efficiency of entry, efficient release of DNA from endosomes, and wide range of cell and tissue targets. Because it consists of only one or two adenovirus proteins instead of the 11 contained in an adenovirus virion and it does not contain the viral genome, it is potentially a safer alternative to recombinant adenovirus.

  4. Acquisition systems for heat transfer measurement

    SciTech Connect

    De Witt, R.J.

    1983-01-01

    Practical heat transfer data acquisition systems are normally characterized by the need for high-resolution, low-drift, low-speed recording devices. Analog devices such as strip chart or circular recorders and FM analog magnetic tape have excellent resolution and work well when data will be presented in temperature versus time format only and need not be processed further. Digital systems are more complex and require an understanding of the following components: digitizing devices, interface bus types, processor requirements, and software design. This paper discusses all the above components of analog and digital data acquisition, as they are used in current practice. Additional information on thermocouple system analysis will aid the user in developing accurate heat transfer measuring systems.

  5. Massive Mitochondrial Gene Transfer in a Parasitic Flowering Plant Clade

    PubMed Central

    Bradley, Robert K.; Sugumaran, M.; Marx, Christopher J.; Rest, Joshua S.; Davis, Charles C.

    2013-01-01

    Recent studies have suggested that plant genomes have undergone potentially rampant horizontal gene transfer (HGT), especially in the mitochondrial genome. Parasitic plants have provided the strongest evidence of HGT, which appears to be facilitated by the intimate physical association between the parasites and their hosts. A recent phylogenomic study demonstrated that in the holoparasite Rafflesia cantleyi (Rafflesiaceae), whose close relatives possess the world's largest flowers, about 2.1% of nuclear gene transcripts were likely acquired from its obligate host. Here, we used next-generation sequencing to obtain the 38 protein-coding and ribosomal RNA genes common to the mitochondrial genomes of angiosperms from R. cantleyi and five additional species, including two of its closest relatives and two host species. Strikingly, our phylogenetic analyses conservatively indicate that 24%–41% of these gene sequences show evidence of HGT in Rafflesiaceae, depending on the species. Most of these transgenic sequences possess intact reading frames and are actively transcribed, indicating that they are potentially functional. Additionally, some of these transgenes maintain synteny with their donor and recipient lineages, suggesting that native genes have likely been displaced via homologous recombination. Our study is the first to comprehensively assess the magnitude of HGT in plants involving a genome (i.e., mitochondria) and a species interaction (i.e., parasitism) where it has been hypothesized to be potentially rampant. Our results establish for the first time that, although the magnitude of HGT involving nuclear genes is appreciable in these parasitic plants, HGT involving mitochondrial genes is substantially higher. This may represent a more general pattern for other parasitic plant clades and perhaps more broadly for angiosperms. PMID:23459037

  6. Massive mitochondrial gene transfer in a parasitic flowering plant clade.

    PubMed

    Xi, Zhenxiang; Wang, Yuguo; Bradley, Robert K; Sugumaran, M; Marx, Christopher J; Rest, Joshua S; Davis, Charles C

    2013-01-01

    Recent studies have suggested that plant genomes have undergone potentially rampant horizontal gene transfer (HGT), especially in the mitochondrial genome. Parasitic plants have provided the strongest evidence of HGT, which appears to be facilitated by the intimate physical association between the parasites and their hosts. A recent phylogenomic study demonstrated that in the holoparasite Rafflesia cantleyi (Rafflesiaceae), whose close relatives possess the world's largest flowers, about 2.1% of nuclear gene transcripts were likely acquired from its obligate host. Here, we used next-generation sequencing to obtain the 38 protein-coding and ribosomal RNA genes common to the mitochondrial genomes of angiosperms from R. cantleyi and five additional species, including two of its closest relatives and two host species. Strikingly, our phylogenetic analyses conservatively indicate that 24%-41% of these gene sequences show evidence of HGT in Rafflesiaceae, depending on the species. Most of these transgenic sequences possess intact reading frames and are actively transcribed, indicating that they are potentially functional. Additionally, some of these transgenes maintain synteny with their donor and recipient lineages, suggesting that native genes have likely been displaced via homologous recombination. Our study is the first to comprehensively assess the magnitude of HGT in plants involving a genome (i.e., mitochondria) and a species interaction (i.e., parasitism) where it has been hypothesized to be potentially rampant. Our results establish for the first time that, although the magnitude of HGT involving nuclear genes is appreciable in these parasitic plants, HGT involving mitochondrial genes is substantially higher. This may represent a more general pattern for other parasitic plant clades and perhaps more broadly for angiosperms.

  7. Parallel Evolution and Horizontal Gene Transfer of the pst Operon in Firmicutes from Oligotrophic Environments

    PubMed Central

    Moreno-Letelier, Alejandra; Olmedo, Gabriela; Eguiarte, Luis E.; Martinez-Castilla, Leon; Souza, Valeria

    2011-01-01

    The high affinity phosphate transport system (pst) is crucial for phosphate uptake in oligotrophic environments. Cuatro Cienegas Basin (CCB) has extremely low P levels and its endemic Bacillus are closely related to oligotrophic marine Firmicutes. Thus, we expected the pst operon of CCB to share the same evolutionary history and protein similarity to marine Firmicutes. Orthologs of the pst operon were searched in 55 genomes of Firmicutes and 13 outgroups. Phylogenetic reconstructions were performed for the pst operon and 14 concatenated housekeeping genes using maximum likelihood methods. Conserved domains and 3D structures of the phosphate-binding protein (PstS) were also analyzed. The pst operon of Firmicutes shows two highly divergent clades with no correlation to the type of habitat nor a phylogenetic congruence, suggesting horizontal gene transfer. Despite sequence divergence, the PstS protein had a similar 3D structure, which could be due to parallel evolution after horizontal gene transfer events. PMID:21461370

  8. Technology insight: gene transfer and the design of novel treatments for rheumatoid arthritis.

    PubMed

    Moritz, Falk; Distler, Oliver; Ospelt, Caroline; Gay, Renate E; Gay, Steffen

    2006-03-01

    Rheumatoid arthritis (RA) is a chronic inflammatory disease characterized by systemic inflammation and joint destruction. Novel therapies have emerged during the past decade, marking a new era in the treatment of RA. Meanwhile, in vivo and in vitro gene-transfer studies have provided valuable insights into mechanisms of disease pathogenesis. Advanced gene-delivery techniques and animal models promise further progress in RA research and the development of novel therapeutic strategies for this disease. In this article we provide an overview of the wide spectrum of potential targets that have been identified so far, discuss currently available gene-transfer methods, and outline the barriers that need to be overcome for these approaches to be successfully applied in daily practice.

  9. Horizontal gene transfer and recombination in Streptococcus dysgalactiae subsp. equisimilis

    PubMed Central

    McNeilly, Celia L.; McMillan, David J.

    2014-01-01

    Streptococcus dysgalactiae subsp. equisimilis (SDSE) is a human pathogen that colonizes the skin or throat, and causes a range of diseases from relatively benign pharyngitis to potentially fatal invasive diseases. While not as virulent as the close relative Streptococcus pyogenes the two share a number of virulence factors and are known to coexist in a human host. Both pre- and post-genomic studies have revealed that horizontal gene transfer (HGT) and recombination occurs between these two organisms and plays a major role in shaping the population structure of SDSE. This review summarizes our current knowledge of HGT and recombination in the evolution of SDSE. PMID:25566202

  10. In vitro study for laser gene transfer in BHK-21 fibroblast cell line

    NASA Astrophysics Data System (ADS)

    Abdel Aziz, M.; Salem, D. S.; Salama, M. S.; Badr, Y.

    2009-02-01

    Modifications to our previously introduced system for laser microbeam cell surgery were carried out in the present work to match animal cells. These modifications included: 1- Using other laser system that used before, Excimer laser with 193 and 308 nm wavelengths. The used laser here, is He-Cd with low power and 441.5 nm wavelength in the visible region. 2- Instead of using pulsed laser, we used here CW He-Cd chopped by electrical chopper, which is synchronized with the mechanical motion of the mobile stage with step 40 microns, according to cell dimensions to avoid puncturing the same cell twice. The advantages of the modified here laser setup for gene transfer is: it is less damaging to the sensitive animal cell which has thin cell membrane. The present work aimed to: 1- Design a modified laser microbeam cell surgery, applicable to animal cells, such as fibroblast cells 2- To examine the efficiency of such system. 3- To assure gene transfer and its expression in the used cells. 4- To evaluate the ultra damages produced from using the laser beam as a modality for gene transfer. On the other wards, to introduce: safe, efficient and less damaging modality for gene transfer in animal cells. To achieve these goals, we applied the introduced here home-made laser setup with its synchronized parameters to introduce pBK-CMV phagemid, containing LacZ and neomycin resistance (neor )genes into BHK-21 fibroblast cell line. The results of the present work showed that: 1- Our modified laser microbeam cell surgery setup proved to be useful and efficient tool for gene transfer into fibroblast cells. 2- The presence and expression of LacZ gene was achieved using histochemical LacZ assay. 3- Selection of G418 antibiotic sensitivity assay confirmed the presence and expression towards stability of neor gene with time. 4- Presence of LacZ and neor genes in the genomic DNA of transfected fibroblast cells was indicated using PCR analysis. 5- Transmission electron microscopy indicated

  11. Forest Resource Information System. Phase 3: System transfer report

    NASA Technical Reports Server (NTRS)

    Mroczynski, R. P. (Principal Investigator)

    1981-01-01

    Transfer of the forest reserve information system (FRIS) from the Laboratory for Applications of Remote Sensing to St. Regis Paper Company is described. Modifications required for the transfer of the LARYS image processing software are discussed. The reformatting, geometric correction, image registration, and documentation performed for preprocessing transfer are described. Data turnaround was improved and geometrically corrected and ground-registered CCT LANDSAT 3 data provided to the user. The technology transfer activities are summarized. An application test performed in order to assess a Florida land acquisition is described. A benefit/cost analysis of FRIS is presented.

  12. Transfer of nonselectable genes into mouse teratocarcinoma cells and transcription of the transferred human. beta. -globin gene

    SciTech Connect

    Wagner, E.F.; Mintz, B.

    1982-02-01

    Teratocarcinoma (TCC) stem cells can function as vehicles for the introduction of specific recombinant genes into mice. Because most genes do not code for a selectable marker, the authors investigated the transformation efficiency of vectors with a linked selectable gene. In one series, TCC cells first selected for thymidine kinase deficiency were treated with DNA from the plasmid vector PtkH..beta..1 containing the human genomic ..beta..-globin gene and the thymidine kinase gene of herpes simplex virus. A high transformation frequency was obtained after selection in hypoxanthine-aminopterin-thymidine medium. Hybridization tests revealed that the majority of transformants had intact copies of the human gene among three to six total copies per cell. These were associated with cellular DNA sequences as judged from the presence of additional new restriction fragments and from stability of the sequences in tumors produced by injecting the cells subcutaneously. Total polyadenylate-containing RNA from cell cultures of two out of four transformants examined showed hybridization to the human gene probe: one RNA species resembled mature human ..beta..-globin mRNA transcripts; the others were of larger size. In differentiating tumors, various tissues, including hematopoietic cells of TCC provenance could be found. In a second model set of experiments, wild-type TCC cells were used to test a dominant-selection scheme with pSV-gpt vectors. Numerous transformants were isolated, and their transfected DNA was apparently stably integrated. Thus, any gene of choice can be transferred into TCC stem cells even without mutagenesis of the cells, and selected cell clones can be characterized. Cells of interest may then be introduced into early embryos to produce new mouse strains with predetermined genetic changes.

  13. Myocardial Gene Transfer: Routes and Devices for Regulation of Transgene Expression by Modulation of Cellular Permeability

    PubMed Central

    Katz, Michael G.; Bridges, Charles R.

    2013-01-01

    Abstract Heart diseases are major causes of morbidity and mortality in Western society. Gene therapy approaches are becoming promising therapeutic modalities to improve underlying molecular processes affecting failing cardiomyocytes. Numerous cardiac clinical gene therapy trials have yet to demonstrate strong positive results and advantages over current pharmacotherapy. The success of gene therapy depends largely on the creation of a reliable and efficient delivery method. The establishment of such a system is determined by its ability to overcome the existing biological barriers, including cellular uptake and intracellular trafficking as well as modulation of cellular permeability. In this article, we describe a variety of physical and mechanical methods, based on the transient disruption of the cell membrane, which are applied in nonviral gene transfer. In addition, we focus on the use of different physiological techniques and devices and pharmacological agents to enhance endothelial permeability. Development of these methods will undoubtedly help solve major problems facing gene therapy. PMID:23427834

  14. Gene Transfer and Molecular Cloning of the Human NGF Receptor

    NASA Astrophysics Data System (ADS)

    Chao, Moses V.; Bothwell, Mark A.; Ross, Alonzo H.; Koprowski, Hilary; Lanahan, Anthony A.; Buck, C. Randall; Sehgal, Amita

    1986-04-01

    Nerve growth factor (NGF) and its receptor are important in the development of cells derived from the neural crest. Mouse L cell transformants have been generated that stably express the human NGF receptor gene transfer with total human DNA. Affinity cross-linking, metabolic labeling and immunoprecipitation, and equilibrium binding with 125I-labeled NGF revealed that this NGF receptor had the same size and binding characteristics as the receptor from human melanoma cells and rat PC12 cells. The sequences encoding the NGF receptor were molecularly cloned using the human Alu repetitive sequence as a probe. A cosmid clone that contained the human NGF receptor gene allowed efficient transfection and expression of the receptor.

  15. Passive immunization against HIV/AIDS by antibody gene transfer.

    PubMed

    Yang, Lili; Wang, Pin

    2014-01-27

    Despite tremendous efforts over the course of many years, the quest for an effective HIV vaccine by the classical method of active immunization remains largely elusive. However, two recent studies in mice and macaques have now demonstrated a new strategy designated as Vectored ImmunoProphylaxis (VIP), which involves passive immunization by viral vector-mediated delivery of genes encoding broadly neutralizing antibodies (bnAbs) for in vivo expression. Robust protection against virus infection was observed in preclinical settings when animals were given VIP to express monoclonal neutralizing antibodies. This unorthodox approach raises new promise for combating the ongoing global HIV pandemic. In this article, we survey the status of antibody gene transfer, review the revolutionary progress on isolation of extremely bnAbs, detail VIP experiments against HIV and its related virus conduced in humanized mice and macaque monkeys, and discuss the pros and cons of VIP and its opportunities and challenges towards clinical applications to control HIV/AIDS endemics.

  16. Laterally Transferred Gene Recruited as a Venom in Parasitoid Wasps.

    PubMed

    Martinson, Ellen O; Martinson, Vincent G; Edwards, Rachel; Mrinalini; Werren, John H

    2016-04-01

    Parasitoid wasps use venom to manipulate the immunity and metabolism of their host insects in a variety of ways to provide resources for their offspring. Yet, how genes are recruited and evolve to perform venom functions remain open questions. A recently recognized source of eukaryotic genome innovation is lateral gene transfer (LGT). Glycoside hydrolase family 19 (GH19) chitinases are widespread in bacteria, microsporidia, and plants where they are used in nutrient acquisition or defense, but have previously not been known in metazoans. In this study, a GH19 chitinase LGT is described from the unicellular microsporidia/Rozella clade into parasitoid wasps of the superfamily Chalcidoidea, where it has become recruited as a venom protein. The GH19 chitinase is present in 15 species of chalcidoid wasps representing four families, and phylogenetic analysis indicates that it was laterally transferred near or before the origin of Chalcidoidea (∼95 Ma). The GH19 chitinase gene is highly expressed in the venom gland of at least seven species, indicating a role in the complex host manipulations performed by parasitoid wasp venom. RNAi knockdown in the model parasitoid Nasonia vitripennis reveals that-following envenomation-the GH19 chitinase induces fly hosts to upregulate genes involved in an immune response to fungi. A second, independent LGT of GH19 chitinase from microsporidia into mosquitoes was also found, also supported by phylogenetic reconstructions. Besides these two LGT events, GH19 chitinase is not found in any other sequenced animal genome, or in any fungi outside the microsporidia/Rozella clade.

  17. Maternal transfer and transcriptional onset of immune genes during ontogenesis in Atlantic cod.

    PubMed

    Seppola, Marit; Johnsen, Hanne; Mennen, Saskia; Myrnes, Bjørnar; Tveiten, Helge

    2009-11-01

    The immune system in teleosts is not completely developed during embryonic and larval stages and immune competence is assumed to be restricted. This study is the first to address whether immune transcripts are maternally transferred to offspring and when immune genes are transcriptionally active in Atlantic cod (Gadus morhua). In unfertilised eggs, transcripts encoding lysozyme and cathelicidin were found indicating maternal transfer of antibacterial transcripts. Lysozyme activity was also present at this stage suggesting the presence of a functional protein. Transcripts of two other putative antibacterial genes (hepcidin and pentraxin) and antiviral genes (ISG15 and LGP2) were absent in unfertilised eggs. The transcriptional onset of these genes occurred during the gastrula period. Transcripts of the heavy chain constant regions of the immunoglobulin (Ig) D, membrane-associated and secreted form of IgM were absent in unfertilised eggs. Transcription of the heavy chain locus commenced at low levels during the segmentation period indicating the onset of B-cell development. Most innate immune genes showed an increase in transcription around hatch and first feeding, indicating a preparation for increased pathogen exposure at this time. Prior to and during metamorphosis all genes showed a pronounced elevation in transcript levels indicating a further maturation of the immune system during this period.

  18. Genome-wide experimental determination of barriers to horizontal gene transfer

    SciTech Connect

    Rubin, Edward; Sorek, Rotem; Zhu, Yiwen; Creevey, Christopher J.; Francino, M. Pilar; Bork, Peer; Rubin, Edward M.

    2007-09-24

    Horizontal gene transfer, in which genetic material is transferred from the genome of one organism to another, has been investigated in microbial species mainly through computational sequence analyses. To address the lack of experimental data, we studied the attempted movement of 246,045 genes from 79 prokaryotic genomes into E. coli and identified genes that consistently fail to transfer. We studied the mechanisms underlying transfer inhibition by placing coding regions from different species under the control of inducible promoters. Their toxicity to the host inhibited transfer regardless of the species of origin and our data suggest that increased gene dosage and associated increased expression is a predominant cause for transfer failure. While these experimental studies examined transfer solely into E. coli, a computational analysis of gene transfer rates across available bacterial and archaeal genomes indicates that the barriers observed in our study are general across the tree of life.

  19. Interspecific evolution: microbial symbiosis, endosymbiosis and gene transfer.

    PubMed

    Hoffmeister, Meike; Martin, William

    2003-08-01

    Microbial symbioses are interesting in their own right and also serve as exemplary models to help biologists to understand two important symbioses in the evolutionary past of eukaryotic cells: the origins of chloroplasts and mitochondria. Most, if not all, microbial symbioses have a chemical basis: compounds produced by one partner are useful for the other. But symbioses can also entail the transfer of genes from one partner to the other, which in some cases cements two cells into a bipartite, co-evolving unit. Here, we discuss some microbial symbioses in which progress is being made in uncovering the nature of symbiotic interactions: anaerobic methane-oxidizing consortia, marine worms that possess endosymbionts instead of a digestive tract, amino acid-producing endosymbionts of aphids, prokaryotic endosymbionts living within a prokaryotic host within mealybugs, endosymbionts of an insect vector of human disease and a photosynthetic sea slug that steals chloroplasts from algae. In the case of chloroplasts and mitochondria, examples of recent and ancient gene transfer to the chromosomes of their host cell illustrate the process of genetic merger in the wake of organelle origins.

  20. Resistance Gene Transfer during Treatments for Experimental Avian Colibacillosis

    PubMed Central

    Dheilly, Alexandra; Le Devendec, Laëtitia; Mourand, Gwenaëlle; Bouder, Axelle; Jouy, Eric

    2012-01-01

    An experiment was conducted in animal facilities to compare the impacts of four avian colibacillosis treatments—oxytetracycline (OTC), trimethoprim-sulfadimethoxine (SXT), amoxicillin (AMX), or enrofloxacin (ENR)—on the susceptibility of Escherichia coli in broiler intestinal tracts. Birds were first orally inoculated with rifampin-resistant E. coli strains bearing plasmid genes conferring resistance to fluoroquinolones (qnr), cephalosporins (blaCTX-M or blaFOX), trimethoprim-sulfonamides, aminoglycosides, or tetracyclines. Feces samples were collected before, during, and after antimicrobial treatments. The susceptibilities of E. coli strains were studied, and resistance gene transfer was analyzed. An increase in the tetracycline-resistant E. coli population was observed only in OTC-treated birds, whereas multiresistant E. coli was detected in the dominant E. coli populations of SXT-, AMX-, or ENR-treated birds. Most multiresistant E. coli strains were susceptible to rifampin and exhibited various pulsed-field gel electrophoresis profiles, suggesting the transfer of one of the multiresistance plasmids from the inoculated strains to other E. coli strains in the intestinal tract. In conclusion, this study clearly illustrates how, in E. coli, “old” antimicrobials may coselect antimicrobial resistance to recent and critical molecules. PMID:21986830

  1. Synthetic Fatty Acids Prevent Plasmid-Mediated Horizontal Gene Transfer

    PubMed Central

    Getino, María; Sanabria-Ríos, David J.; Fernández-López, Raúl; Campos-Gómez, Javier; Sánchez-López, José M.; Fernández, Antonio; Carballeira, Néstor M.

    2015-01-01

    ABSTRACT Bacterial conjugation constitutes a major horizontal gene transfer mechanism for the dissemination of antibiotic resistance genes among human pathogens. Antibiotic resistance spread could be halted or diminished by molecules that interfere with the conjugation process. In this work, synthetic 2-alkynoic fatty acids were identified as a novel class of conjugation inhibitors. Their chemical properties were investigated by using the prototype 2-hexadecynoic acid and its derivatives. Essential features of effective inhibitors were the carboxylic group, an optimal long aliphatic chain of 16 carbon atoms, and one unsaturation. Chemical modification of these groups led to inactive or less-active derivatives. Conjugation inhibitors were found to act on the donor cell, affecting a wide number of pathogenic bacterial hosts, including Escherichia, Salmonella, Pseudomonas, and Acinetobacter spp. Conjugation inhibitors were active in inhibiting transfer of IncF, IncW, and IncH plasmids, moderately active against IncI, IncL/M, and IncX plasmids, and inactive against IncP and IncN plasmids. Importantly, the use of 2-hexadecynoic acid avoided the spread of a derepressed IncF plasmid into a recipient population, demonstrating the feasibility of abolishing the dissemination of antimicrobial resistances by blocking bacterial conjugation. PMID:26330514

  2. Intra- and inter-generic transfer of pathogenicity island-encoded virulence genes by cos phages.

    PubMed

    Chen, John; Carpena, Nuria; Quiles-Puchalt, Nuria; Ram, Geeta; Novick, Richard P; Penadés, José R

    2015-05-01

    Bacteriophage-mediated horizontal gene transfer is one of the primary driving forces of bacterial evolution. The pac-type phages are generally thought to facilitate most of the phage-mediated gene transfer between closely related bacteria, including that of mobile genetic elements-encoded virulence genes. In this study, we report that staphylococcal cos-type phages transferred the Staphylococcus aureus pathogenicity island SaPIbov5 to non-aureus staphylococcal species and also to different genera. Our results describe the first intra- and intergeneric transfer of a pathogenicity island by a cos phage, and highlight a gene transfer mechanism that may have important implications for pathogen evolution.

  3. Block Transfer: A Private Career College Student's Ladder into the BC Transfer System. Special Report

    ERIC Educational Resources Information Center

    McIvor, Raili

    2010-01-01

    In June 2008 the British Columbia Council on Admissions and Transfer (BCCAT) approved a new policy to publish in the BC Transfer Guide block transfer agreements (BTAs) between BC Transfer System member institutions and private post-secondary institutions accredited by the Private Career Training Institutions Agency (PCTIA). This new policy was…

  4. Localized gene transfer and its application for the study of central cardiovascular control.

    PubMed

    Hirooka, Yoshitaka

    2006-06-30

    The arterial baroreceptor reflex is the major feedback control system that acts to stabilize blood pressure. Abnormalities of this reflex are considered to be an underlying mechanism in the cardiovascular diseases such as hypertension and heart failure. There is accumulating evidence, however, that central nervous system mechanisms are involved in the enhanced sympathetic drive that occurs in these disease states. This article reviews studies performed in our laboratory in which a gene transfer technique, in combination with other methods, was used to determine the functional role of the central control of cardiovascular regulation. We developed a technique to transfer adenovirus vectors encoding specific genes into the nucleus tractus solitarii (NTS) or the rostral ventral medulla (RVLM) of rats in vivo. We applied this technique to hypertensive rats as well as in mice with heart failure to explore the pathophysiological significance of nitric oxide, reactive oxygen species, and Rho-kinase.

  5. Simultaneous gene quantitation of multiple genes in individual bovine nuclear transfer blastocysts.

    PubMed

    Smith, Craig; Berg, Debbie; Beaumont, Sue; Standley, Neil T; Wells, David N; Pfeffer, Peter L

    2007-01-01

    During somatic cell nuclear transfer (NT), the transcriptional status of the donor cell has to be reprogrammed to reflect that of an embryo. We analysed the accuracy of this process by comparing transcript levels of four developmentally important genes (Oct4, Otx2, Ifitm3, GATA6), a gene involved in epigenetic regulation (Dnmt3a) and three housekeeping genes (beta-actin, beta-tubulin and GAPDH) in 21 NT blastocysts with that in genetically half-identical in vitro produced (IVP, n=19) and in vivo (n=15) bovine embryos. We have optimised an RNA-isolation and SYBR-green-based real-time RT-PCR procedure allowing the reproducible absolute quantification of multiple genes from a single blastocyst. Our data indicated that transcript levels did not differ significantly between stage and grade-matched zona-free NT and IVP embryos except for Ifitm3/Fragilis, which was expressed at twofold higher levels in NT blastocysts. Ifitm3 expression is confined to the inner cell mass at day 7 blastocysts and to the epiblast in day 14 embryos. No ectopic expression in the trophectoderm was seen in NT embryos. Gene expression in NT and IVP embryos increased between two- and threefold for all eight genes from early to late blastocyst stages. This increase exceeded the increase in cell number over this time period indicating an increase in transcript number per cell. Embryo quality (morphological grading) was correlated to cell number for NT and IVP embryos with grade 3 blastocysts containing 30% fewer cells. However, only NT embryos displayed a significant reduction in gene expression (50%) with loss of quality. Variability in gene expression levels was not significantly different in NT, IVP or in vivo embryos but differed among genes, suggesting that the stringency of regulation is intrinsic to a gene and not affected by culture or nuclear transfer. Oct4 levels exhibited the lowest variability. Analysing the total variability of all eight genes for individual embryos revealed that in

  6. Ultrasound-mediated gene transfer (sonoporation) in fibrin-based matrices: potential for use in tissue regeneration.

    PubMed

    Nomikou, Nikolitsa; Feichtinger, Georg A; Redl, Heinz; McHale, Anthony P

    2016-01-01

    It has been suggested that gene transfer into donor cells is an efficient and practical means of locally supplying requisite growth factors for applications in tissue regeneration. Here we describe, for the first time, an ultrasound-mediated system that can non-invasively facilitate gene transfer into cells entrapped within fibrin-based matrices. Since ultrasound-mediated gene transfer is enhanced using microbubbles, we compared the efficacy of neutral and cationic forms of these reagents on the ultrasound-stimulated gene transfer process in gel matrices. In doing so we demonstrated the beneficial effects associated with the use of cationic microbubble preparations that interact directly with cells and nucleic acid within matrices. In some cases, gene expression was increased two-fold in gel matrices when cationic microbubbles were compared with neutral microbubbles. In addition, incorporating collagen into fibrin gels yielded a 25-fold increase in gene expression after application of ultrasound to microbubble-containing matrices. We suggest that this novel system may facilitate non-invasive temporal and spatial control of gene transfer in gel-based matrices for the purposes of tissue regeneration.

  7. Lateral Gene Transfer and Gene Duplication Played a Key Role in the Evolution of Mastigamoeba balamuthi Hydrogenosomes

    PubMed Central

    Nývltová, Eva; Stairs, Courtney W.; Hrdý, Ivan; Rídl, Jakub; Mach, Jan; Pačes, Jan; Roger, Andrew J.; Tachezy, Jan

    2015-01-01

    Lateral gene transfer (LGT) is an important mechanism of evolution for protists adapting to oxygen-poor environments. Specifically, modifications of energy metabolism in anaerobic forms of mitochondria (e.g., hydrogenosomes) are likely to have been associated with gene transfer from prokaryotes. An interesting question is whether the products of transferred genes were directly targeted into the ancestral organelle or initially operated in the cytosol and subsequently acquired organelle-targeting sequences. Here, we identified key enzymes of hydrogenosomal metabolism in the free-living anaerobic amoebozoan Mastigamoeba balamuthi and analyzed their cellular localizations, enzymatic activities, and evolutionary histories. Additionally, we characterized 1) several canonical mitochondrial components including respiratory complex II and the glycine cleavage system, 2) enzymes associated with anaerobic energy metabolism, including an unusual D-lactate dehydrogenase and acetyl CoA synthase, and 3) a sulfate activation pathway. Intriguingly, components of anaerobic energy metabolism are present in at least two gene copies. For each component, one copy possesses an mitochondrial targeting sequence (MTS), whereas the other lacks an MTS, yielding parallel cytosolic and hydrogenosomal extended glycolysis pathways. Experimentally, we confirmed that the organelle targeting of several proteins is fully dependent on the MTS. Phylogenetic analysis of all extended glycolysis components suggested that these components were acquired by LGT. We propose that the transformation from an ancestral organelle to a hydrogenosome in the M. balamuthi lineage involved the lateral acquisition of genes encoding extended glycolysis enzymes that initially operated in the cytosol and that established a parallel hydrogenosomal pathway after gene duplication and MTS acquisition. PMID:25573905

  8. Intraspecies Transfer of the Chromosomal Acinetobacter baumannii blaNDM-1 Carbapenemase Gene

    PubMed Central

    Krahn, Thomas; Wibberg, Daniel; Maus, Irena; Winkler, Anika; Bontron, Séverine; Sczyrba, Alexander; Nordmann, Patrice; Pühler, Alfred; Poirel, Laurent

    2016-01-01

    The species Acinetobacter baumannii is one of the most important multidrug-resistant human pathogens. To determine its virulence and antibiotic resistance determinants, the genome of the nosocomial blaNDM-1-positive A. baumannii strain R2090 originating from Egypt was completely sequenced. Genome analysis revealed that strain R2090 is highly related to the community-acquired Australian A. baumannii strain D1279779. The two strains belong to sequence type 267 (ST267). Isolate R2090 harbored the chromosomally integrated transposon Tn125 carrying the carbapenemase gene blaNDM-1 that is not present in the D1279779 genome. To test the transferability of the metallo-β-lactamase (MBL) gene region, the clinical isolate R2090 was mated with the susceptible A. baumannii recipient CIP 70.10, and the carbapenem-resistant derivative R2091 was obtained. Genome sequencing of the R2091 derivative revealed that it had received an approximately 66-kb region comprising the transposon Tn125 embedding the blaNDM-1 gene. This region had integrated into the chromosome of the recipient strain CIP 70.10. From the four known mechanisms for horizontal gene transfer (conjugation, outer membrane vesicle-mediated transfer, transformation, and transduction), conjugation could be ruled out, since strain R2090 lacks any plasmid, and a type IV secretion system is not encoded in its chromosome. However, strain R2090 possesses three putative prophages, two of which were predicted to be complete and therefore functional. Accordingly, it was supposed that the transfer of the resistance gene region from the clinical isolate R2090 to the recipient occurred by general transduction facilitated by one of the prophages present in the R2090 genome. Hence, phage-mediated transduction has to be taken into account for the dissemination of antibiotic resistance genes within the species A. baumannii. PMID:26953198

  9. Adaptive Horizontal Gene Transfers between Multiple Cheese-Associated Fungi

    PubMed Central

    Ropars, Jeanne; Rodríguez de la Vega, Ricardo C.; López-Villavicencio, Manuela; Gouzy, Jérôme; Sallet, Erika; Dumas, Émilie; Lacoste, Sandrine; Debuchy, Robert; Dupont, Joëlle; Branca, Antoine; Giraud, Tatiana

    2015-01-01

    Summary Domestication is an excellent model for studies of adaptation because it involves recent and strong selection on a few, identified traits [1–5]. Few studies have focused on the domestication of fungi, with notable exceptions [6–11], despite their importance to bioindustry [12] and to a general understanding of adaptation in eukaryotes [5]. Penicillium fungi are ubiquitous molds among which two distantly related species have been independently selected for cheese making—P. roqueforti for blue cheeses like Roquefort and P. camemberti for soft cheeses like Camembert. The selected traits include morphology, aromatic profile, lipolytic and proteolytic activities, and ability to grow at low temperatures, in a matrix containing bacterial and fungal competitors [13–15]. By comparing the genomes of ten Penicillium species, we show that adaptation to cheese was associated with multiple recent horizontal transfers of large genomic regions carrying crucial metabolic genes. We identified seven horizontally transferred regions (HTRs) spanning more than 10 kb each, flanked by specific transposable elements, and displaying nearly 100% identity between distant Penicillium species. Two HTRs carried genes with functions involved in the utilization of cheese nutrients or competition and were found nearly identical in multiple strains and species of cheese-associated Penicillium fungi, indicating recent selective sweeps; they were experimentally associated with faster growth and greater competitiveness on cheese and contained genes highly expressed in the early stage of cheese maturation. These findings have industrial and food safety implications and improve our understanding of the processes of adaptation to rapid environmental changes. PMID:26412136

  10. The efficiency of genetic transformation of mammalian cells by transfection and microinjection depends on the transferred gene.

    PubMed

    Strauss, M; Kiessling, U; Zavision, B A; Povitza, O N; Tikhonenko, T I; Geissler, E

    1983-01-01

    The efficiency of genetic transformation of mammalian cells was analysed with respect to the kind of the transferred gene and the selective system. Plasmids pAGO and pAG60 harboring the thymidine kinase gene of Herpes simplex virus type 1 and the bacterial neomycin resistance gene, respectively, were compared concerning their ability to transform mouse Ltk-aprt- cells. Using the calcium phosphate technique the neomycin resistance gene transformed at least ten times more efficiently than the thymidine kinase gene (3 X 10(-3) versus 2 X 10(-4] whereas the difference is even more impressive following microinjection of the plasmids into the nuclei (2 X 10(-1) versus 2.5 X 10(-3]. The neomycin system also proved to be more effective in secondary gene transfer experiments and, thus, seems to be the most convenient marker for cotransfer experiments.

  11. Comparative analysis of magnetosome gene clusters in magnetotactic bacteria provides further evidence for horizontal gene transfer.

    PubMed

    Jogler, Christian; Kube, Michael; Schübbe, Sabrina; Ullrich, Susanne; Teeling, Hanno; Bazylinski, Dennis A; Reinhardt, Richard; Schüler, Dirk

    2009-05-01

    The organization of magnetosome genes was analysed in all available complete or partial genomic sequences of magnetotactic bacteria (MTB), including the magnetosome island (MAI) of the magnetotactic marine vibrio strain MV-1 determined in this study. The MAI was found to differ in gene content and organization between Magnetospirillum species and strains MV-1 or MC-1. Although a similar organization of magnetosome genes was found in all MTB, distinct variations in gene order and sequence similarity were uncovered that may account for the observed diversity of biomineralization, cell biology and magnetotaxis found in various MTB. While several magnetosome genes were present in all MTB, others were confined to Magnetospirillum species, indicating that the minimal set of genes required for magnetosome biomineralization might be smaller than previously suggested. A number of novel candidate genes were implicated in magnetosome formation by gene cluster comparison. Based on phylogenetic and compositional evidence we present a model for the evolution of magnetotaxis within the Alphaproteobacteria, which suggests the independent horizontal transfer of magnetosome genes from an unknown ancestor of magnetospirilla into strains MC-1 and MV-1.

  12. Immunological ignorance allows long-term gene expression after perinatal recombinant adeno-associated virus-mediated gene transfer to murine airways.

    PubMed

    Carlon, Marianne S; Vidović, Dragana; Dooley, James; da Cunha, Marina Mori; Maris, Michael; Lampi, Youlia; Toelen, Jaan; Van den Haute, Chris; Baekelandt, Veerle; Deprest, Jan; Verbeken, Erik; Liston, Adrian; Gijsbers, Rik; Debyser, Zeger

    2014-06-01

    Gene therapy of the lung has the potential to treat life-threatening diseases such as cystic fibrosis and α(1)-antitrypsin or surfactant deficiencies. A major hurdle for successful gene therapy is the development of an immune response against the transgene and/or viral vector. We hypothesized that by targeting the airways in the perinatal period, induction of an immune response against the vector particle could be prevented because of immaturity of the immune system, in turn allowing repeated gene transfer later in adult life to ensure long-term gene expression. Therefore, we readministered recombinant adeno-associated viral vector serotype 5 (rAAV2/5) to mouse airways 3 and 6 months after initial perinatal gene transfer. Our findings demonstrate that perinatal rAAV2/5-mediated gene transfer to the airways avoids a strong immune response. This immunological ignorance allows the readministration of an autologous vector later in adult life, resulting in efficient and stable gene transfer up to 7 months, without evidence of a decrease in transgene expression. Together, these data provide a basis to further explore perinatal gene therapy for pulmonary conditions with adequate gene expression up to 7 months.

  13. Ultrasound Gene Transfer into Fibroblast Cells using Microbubbles

    NASA Astrophysics Data System (ADS)

    Nakamura, Yoji; Hirayama, Kota; Yoshinaka, Kiyoshi; Tei, Yuichi; Takagi, Shu; Matsumoto, Yoichiro

    2009-04-01

    Ultrasound is widely applied in the medical field and offers the strong advantages of non-invasiveness and high-selectivity. Gene transfer using ultrasound, which is called sonoporation, is one application. Ultrasound has the potential to deliver therapeutic materials such as genes, drugs or proteins into cells. Microbubbles are known to be able to improve delivery efficiency. This is attributed to therapeutic materials passing through the cell membrane after permeability is increased by destruction or oscillation of microbubbles. The present study tried to deliver the GFP plasmids into fibroblast cells. Cells were cultured in 6-well culture plates and exposed to ultrasound (frequency, 2.1 MHz; wave pattern, duty cycle 10%; intensity, 0-26 W/cm2; time, 0-200 s) transmitted through medium containing microbubbles (Levovist® (void fraction, 8×10-5) or Sonazoid® (void fraction, 0-24×10-4)) and GFP plasmids at a concentration of 15 μg/mL. Density of microbubbles after ultrasound irradiation was measured. When ultrasound intensity was increased with Levovist® 8×10-4, transfection efficiency increased, cell viability decreased and microbubbles disappeared. With Sonazoid®, transfection efficiency and cell viability were basically unchanged and microbubbles decreased, but did not disappear. Transfection efficiency also improved with increased ultrasound irradiation time or microbubble density. Microbubble destruction appeared to have the main effect on gene transfection under Levovist® and microbubble oscillation had the main effect under Sonazoid®.

  14. Phylogenetic analyses of cyanobacterial genomes: Quantification of horizontal gene transfer events

    PubMed Central

    Zhaxybayeva, Olga; Gogarten, J. Peter; Charlebois, Robert L.; Doolittle, W. Ford; Papke, R. Thane

    2006-01-01

    Using 1128 protein-coding gene families from 11 completely sequenced cyanobacterial genomes, we attempt to quantify horizontal gene transfer events within cyanobacteria, as well as between cyanobacteria and other phyla. A novel method of detecting and enumerating potential horizontal gene transfer events within a group of organisms based on analyses of “embedded quartets” allows us to identify phylogenetic signal consistent with a plurality of gene families, as well as to delineate cases of conflict to the plurality signal, which include horizontally transferred genes. To infer horizontal gene transfer events between cyanobacteria and other phyla, we added homologs from 168 available genomes. We screened phylogenetic trees reconstructed for each of these extended gene families for highly supported monophyly of cyanobacteria (or lack of it). Cyanobacterial genomes reveal a complex evolutionary history, which cannot be represented by a single strictly bifurcating tree for all genes or even most genes, although a single completely resolved phylogeny was recovered from the quartets’ plurality signals. We find more conflicts within cyanobacteria than between cyanobacteria and other phyla. We also find that genes from all functional categories are subject to transfer. However, in interphylum as compared to intraphylum transfers, the proportion of metabolic (operational) gene transfers increases, while the proportion of informational gene transfers decreases. PMID:16899658

  15. Advanced orbit transfer vehicle propulsion system study

    NASA Technical Reports Server (NTRS)

    Cathcart, J. A.; Cooper, T. W.; Corringrato, R. M.; Cronau, S. T.; Forgie, S. C.; Harder, M. J.; Mcallister, J. G.; Rudman, T. J.; Stoneback, V. W.

    1985-01-01

    A reuseable orbit transfer vehicle concept was defined and subsequent recommendations for the design criteria of an advanced LO2/LH2 engine were presented. The major characteristics of the vehicle preliminary design include a low lift to drag aerocapture capability, main propulsion system failure criteria of fail operational/fail safe, and either two main engines with an attitude control system for backup or three main engines to meet the failure criteria. A maintenance and servicing approach was also established for the advanced vehicle and engine concepts. Design tradeoff study conclusions were based on the consideration of reliability, performance, life cycle costs, and mission flexibility.

  16. Supra-operonic clusters of functionally related genes (SOCs) are a source of horizontal gene co-transfers

    PubMed Central

    Pang, Tin Yau; Lercher, Martin J.

    2017-01-01

    Adaptation of bacteria occurs predominantly via horizontal gene transfer (HGT). While it is widely recognized that horizontal acquisitions frequently encompass multiple genes, it is unclear what the size distribution of successfully transferred DNA segments looks like and what evolutionary forces shape this distribution. Here, we identified 1790 gene family pairs that were consistently co-gained on the same branches across a phylogeny of 53 E. coli strains. We estimated a lower limit of their genomic distances at the time they were transferred to their host genomes; this distribution shows a sharp upper bound at 30 kb. The same gene-pairs can have larger distances (up to 70 kb) in other genomes. These more distant pairs likely represent recent acquisitions via transduction that involve the co-transfer of excised prophage genes, as they are almost always associated with intervening phage-associated genes. The observed distribution of genomic distances of co-transferred genes is much broader than expected from a model based on the co-transfer of genes within operons; instead, this distribution is highly consistent with the size distribution of supra-operonic clusters (SOCs), groups of co-occurring and co-functioning genes that extend beyond operons. Thus, we propose that SOCs form a basic unit of horizontal gene transfer. PMID:28067311

  17. Long range inductive power transfer system

    NASA Astrophysics Data System (ADS)

    Lawson, James; Pinuela, Manuel; Yates, David C.; Lucyszyn, Stepan; Mitcheson, Paul D.

    2013-12-01

    We report upon a recently developed long range inductive power transfer system (IPT) designed to power remote sensors with mW level power consumption at distances up to 7 m. In this paper an inductive link is established between a large planar (1 × 1 m) transmit coil (Tx) and a small planer (170 × 170 mm) receiver coil (Rx), demonstrating the viability of highly asymmetrical coil configurations that real-world applications such as sensor networks impose. High Q factor Tx and Rx coils required for viable power transfer efficiencies over such distances are measured using a resonant method. The applicability of the Class-E amplifier in very low magnetic coupling scenarios and at the high frequencies of operation required for high Q operation is demonstrated by its usage as the Tx coil driver.

  18. Air Storage System Energy Transfer (ASSET) plants

    NASA Astrophysics Data System (ADS)

    Stys, Z. S.

    1983-09-01

    The design features and performance capabilities of Air Storage System Energy Transfer (ASSET) plants for transferring off-peak utility electricity to on-peak hours are described. The plant operations involve compressing ambient air with an axial flow compressor and depositing it in an underground reservoir at 70 bar pressure. Released during a peaking cycle, the pressure is reduced to 43 bar, the air is heated to 550 C, passed through an expander after a turbine, and passed through a low pressure combustion chamber to be heated to 850 C. A West German plant built in 1978 to supply over 300 MW continuous power for up to two hours is detailed, noting its availability factor of nearly 98 percent and power delivery cost of $230/kW installed. A plant being constructed in Illinois will use limestone caverns as the air storage tank.

  19. In vivo gene transfer targeting in pancreatic adenocarcinoma with cell surface antigens

    PubMed Central

    2012-01-01

    Background Pancreatic ductal adenocarcinoma is a deadly malignancy resistant to current therapies. It is critical to test new strategies, including tumor-targeted delivery of therapeutic agents. This study tested the possibility to target the transfer of a suicide gene in tumor cells using an oncotropic lentiviral vector. Results Three cell surface markers were evaluated to target the transduction of cells by lentiviruses pseudotyped with a modified glycoprotein from Sindbis virus. Only Mucin-4 and the Claudin-18 proteins were found efficient for targeted lentivirus transductions in vitro. In subcutaneous xenografts of human pancreatic cancer cells models, Claudin-18 failed to achieve efficient gene transfer but Mucin-4 was found very potent. Human pancreatic tumor cells were modified to express a fluorescent protein detectable in live animals by bioimaging, to perform a direct non invasive and costless follow up of the tumor growth. Targeted gene transfer of a bicistronic transgene bearing a luciferase gene and the herpes simplex virus thymidine kinase gene into orthotopic grafts was carried out with Mucin-4 oncotropic lentiviruses. By contrast to the broad tropism VSV-G carrying lentivirus, this oncotropic lentivirus was found to transduce specifically tumor cells, sparing normal pancreatic cells in vivo. Transduced cells disappeared after ganciclovir treatment while the orthotopic tumor growth was slowed down. Conclusion This work considered for the first time three aspect of pancreatic adenocarcinoma targeted therapy. First, lentiviral transduction of human pancreatic tumor cells was possible when cells were grafted orthotopically. Second, we used a system targeting the tumor cells with cell surface antigens and sparing the normal cells. Finally, the TK/GCV anticancer system showed promising results in vivo. Importantly, the approach presented here appeared to be a safer, much more specific and an as efficient way to perform gene delivery in pancreatic tumors

  20. Horizontal gene transfer and gene dosage drives adaptation to wood colonization in a tree pathogen

    DOE PAGES

    Dhillon, Braham; Feau, Nicolas; Aerts, Andrea L.; ...

    2015-03-02

    Some of the most damaging tree diseases are caused by pathogens that induce cankers, a stem deformation often lethal. To investigate the cause of this adaptation, we sequenced the genomes of poplar pathogens that do and do not cause cankers. We found a unique cluster of genes that produce secondary metabolites and are co-activated when the canker pathogen is grown on poplar wood and leaves. The gene genealogy is discordant with the species phylogeny, showing a signature of horizontal transfer from fungi associated with wood decay. Furthermore, genes encoding hemicellulose-degrading enzymes are up-regulated on poplar wood chips, with some havingmore » been acquired horizontally. In conclusion, we propose that adaptation to colonize poplar woody stems is the result of acquisition of these genes.« less

  1. Horizontal gene transfer and gene dosage drives adaptation to wood colonization in a tree pathogen

    SciTech Connect

    Dhillon, Braham; Feau, Nicolas; Aerts, Andrea L.; Beauseigle, Stéphanie; Bernier, Louis; Copeland, Alex; Foster, Adam; Gill, Navdeep; Henrissat, Bernard; Herath, Padmini; LaButti, Kurt M.; Levasseur, Anthony; Lindquist, Erika A.; Majoor, Eline; Ohm, Robin A.; Pangilinan, Jasmyn L.; Pribowo, Amadeus; Saddler, John N.; Sakalidis, Monique L.; de Vries, Ronald P.; Grigoriev, Igor V.; Goodwin, Stephen B.; Tanguay, Philippe; Hamelin, Richard C.

    2015-03-02

    Some of the most damaging tree diseases are caused by pathogens that induce cankers, a stem deformation often lethal. To investigate the cause of this adaptation, we sequenced the genomes of poplar pathogens that do and do not cause cankers. We found a unique cluster of genes that produce secondary metabolites and are co-activated when the canker pathogen is grown on poplar wood and leaves. The gene genealogy is discordant with the species phylogeny, showing a signature of horizontal transfer from fungi associated with wood decay. Furthermore, genes encoding hemicellulose-degrading enzymes are up-regulated on poplar wood chips, with some having been acquired horizontally. In conclusion, we propose that adaptation to colonize poplar woody stems is the result of acquisition of these genes.

  2. Horizontal gene transfer: building the web of life.

    PubMed

    Soucy, Shannon M; Huang, Jinling; Gogarten, Johann Peter

    2015-08-01

    Horizontal gene transfer (HGT) is the sharing of genetic material between organisms that are not in a parent-offspring relationship. HGT is a widely recognized mechanism for adaptation in bacteria and archaea. Microbial antibiotic resistance and pathogenicity are often associated with HGT, but the scope of HGT extends far beyond disease-causing organisms. In this Review, we describe how HGT has shaped the web of life using examples of HGT among prokaryotes, between prokaryotes and eukaryotes, and even between multicellular eukaryotes. We discuss replacement and additive HGT, the proposed mechanisms of HGT, selective forces that influence HGT, and the evolutionary impact of HGT on ancestral populations and existing populations such as the human microbiome.

  3. Studies of DEAE-dextran-mediated gene transfer.

    PubMed

    Yang, Y W; Yang, J C

    1997-02-01

    DEAE-dextran-mediated gene transfer was studied for the introduction of pSV2neo DNA into Fisher-rat 3T3 (FR3T3) cells. Zeta (zeta) potentials of the DEAE-dextran-DNA complexes and FR3T3 cells were found to be dependent on the concentration of DEAE-dextran in the medium. The maximum transfection efficiency occurred at a DEAE-dextran/DNA ratio of 50:1 or thereabouts. The interaction between DNA and cells is determined by the adsorption process. The results obtained, along with the correlation between the kinetic adsorption behaviour of 3H-labelled DNA and the transfection efficiency, indicated that adsorption of DEAE-dextran-DNA complexes to the negatively charged cell surfaces, due to electrostatic and dispersion attraction, plays the decisive role in determining the DNA transfection efficiencies.

  4. Statistical Mechanics of Horizontal Gene Transfer in Evolutionary Ecology

    NASA Astrophysics Data System (ADS)

    Chia, Nicholas; Goldenfeld, Nigel

    2011-04-01

    The biological world, especially its majority microbial component, is strongly interacting and may be dominated by collective effects. In this review, we provide a brief introduction for statistical physicists of the way in which living cells communicate genetically through transferred genes, as well as the ways in which they can reorganize their genomes in response to environmental pressure. We discuss how genome evolution can be thought of as related to the physical phenomenon of annealing, and describe the sense in which genomes can be said to exhibit an analogue of information entropy. As a direct application of these ideas, we analyze the variation with ocean depth of transposons in marine microbial genomes, predicting trends that are consistent with recent observations using metagenomic surveys.

  5. Extensive Intra-Kingdom Horizontal Gene Transfer Converging on a Fungal Fructose Transporter Gene

    PubMed Central

    Coelho, Marco A.; Gonçalves, Carla; Sampaio, José Paulo; Gonçalves, Paula

    2013-01-01

    Comparative genomics revealed in the last decade a scenario of rampant horizontal gene transfer (HGT) among prokaryotes, but for fungi a clearly dominant pattern of vertical inheritance still stands, punctuated however by an increasing number of exceptions. In the present work, we studied the phylogenetic distribution and pattern of inheritance of a fungal gene encoding a fructose transporter (FSY1) with unique substrate selectivity. 109 FSY1 homologues were identified in two sub-phyla of the Ascomycota, in a survey that included 241 available fungal genomes. At least 10 independent inter-species instances of horizontal gene transfer (HGT) involving FSY1 were identified, supported by strong phylogenetic evidence and synteny analyses. The acquisition of FSY1 through HGT was sometimes suggestive of xenolog gene displacement, but several cases of pseudoparalogy were also uncovered. Moreover, evidence was found for successive HGT events, possibly including those responsible for transmission of the gene among yeast lineages. These occurrences do not seem to be driven by functional diversification of the Fsy1 proteins because Fsy1 homologues from widely distant lineages, including at least one acquired by HGT, appear to have similar biochemical properties. In summary, retracing the evolutionary path of the FSY1 gene brought to light an unparalleled number of independent HGT events involving a single fungal gene. We propose that the turbulent evolutionary history of the gene may be linked to the unique biochemical properties of the encoded transporter, whose predictable effect on fitness may be highly variable. In general, our results support the most recent views suggesting that inter-species HGT may have contributed much more substantially to shape fungal genomes than heretofore assumed. PMID:23818872

  6. Submersible pumping system with heat transfer mechanism

    DOEpatents

    Hunt, Daniel Francis Alan; Prenger, F. Coyne; Hill, Dallas D; Jankowski, Todd Andrew

    2014-04-15

    A submersible pumping system for downhole use in extracting fluids containing hydrocarbons from a well. In one embodiment, the pumping system comprises a rotary induction motor, a motor casing, one or more pump stages, and a cooling system. The rotary induction motor rotates a shaft about a longitudinal axis of rotation. The motor casing houses the rotary induction motor such that the rotary induction motor is held in fluid isolation from the fluid being extracted. The pump stages are attached to the shaft outside of the motor casing, and are configured to impart fluid being extracted from the well with an increased pressure. The cooling system is disposed at least partially within the motor casing, and transfers heat generated by operation of the rotary induction motor out of the motor casing.

  7. Applying horizontal gene transfer phenomena to enhance non-viral gene therapy

    PubMed Central

    Elmer, Jacob J.; Christensen, Matthew D.; Rege, Kaushal

    2014-01-01

    Horizontal gene transfer (HGT) is widespread amongst prokaryotes, but eukaryotes tend to be far less promiscuous with their genetic information. However, several examples of HGT from pathogens into eukaryotic cells have been discovered and mimicked to improve non-viral gene delivery techniques. For example, several viral proteins and DNA sequences have been used to significantly increase cytoplasmic and nuclear gene delivery. Plant genetic engineering is routinely performed with the pathogenic bacterium Agrobacterium tumefaciens and similar pathogens (e.g. Bartonella henselae) may also be able to transform human cells. Intracellular parasites like Trypanosoma cruzi may also provide new insights into overcoming cellular barriers to gene delivery. Finally, intercellular nucleic acid transfer between host cells will also be briefly discussed. This article will review the unique characteristics of several different viruses and microbes and discuss how their traits have been successfully applied to improve non-viral gene delivery techniques. Consequently, pathogenic traits that originally caused diseases may eventually be used to treat many genetic diseases. PMID:23994344

  8. Expertise transfer for expert system design

    SciTech Connect

    Boose, J.H.

    1986-01-01

    This book is about the Expertise Transfer System-a computer program which interviews experts and helps them build expert systems, i.e. computer programs that use knowledge from experts to make decisions and judgements under conditions of uncertainty. The techniques are useful to anyone who uses decision-making information based on the expertise of others. The methods can also be applied to personal decision-making. The interviewing methodology is borrowed from a branch of psychology called Personal Construct Theory. It is not necessary to use a computer to take advantage of the techniques from Personal Construction Theory; the fundamental procedures used by the Expertise Transfer System can be performed using paper and pencil. It is not necessary that the reader understand very much about computers to understand the ideas in this book. The few relevant concepts from computer science and expert systems that are needed are explained in a straightforward manner. Ideas from Personal Construct Psychology are also introduced as needed.

  9. Oxygen Transfer Characteristics of Miniaturized Bioreactor Systems

    PubMed Central

    Kirk, Timothy V; Szita, Nicolas

    2013-01-01

    Since their introduction in 2001 miniaturized bioreactor systems have made great advances in function and performance. In this article the dissolved oxygen (DO) transfer performance of submilliliter microbioreactors, and 1–10 mL minibioreactors was examined. Microbioreactors have reached kLa values of 460 h-1, and are offering instrumentation and some functionality comparable to production systems, but at high throughput screening volumes. Minibioreactors, aside from one 1,440 h-1 kLa system, have not offered as high rates of DO transfer, but have demonstrated superior integration with automated fluid handling systems. Microbioreactors have been typically limited to studies with E. coli, while minibioreactors have offered greater versatility in this regard. Further, mathematical relationships confirming the applicability of kLa measurements across all scales have been derived, and alternatives to fluorescence lifetime DO sensors have been evaluated. Finally, the influence on reactor performance of oxygen uptake rate (OUR), and the possibility of its real-time measurement have been explored. Biotechnol. Bioeng. 2013; 110: 1005–1019. © 2012 Wiley Periodicals, Inc. PMID:23280578

  10. Widespread impact of horizontal gene transfer on plant colonization of land

    PubMed Central

    Yue, Jipei; Hu, Xiangyang; Sun, Hang; Yang, Yongping; Huang, Jinling

    2012-01-01

    In complex multicellular eukaryotes such as animals and plants, horizontal gene transfer is commonly considered rare with very limited evolutionary significance. Here we show that horizontal gene transfer is a dynamic process occurring frequently in the early evolution of land plants. Our genome analyses of the moss Physcomitrella patens identified 57 families of nuclear genes that were acquired from prokaryotes, fungi or viruses. Many of these gene families were transferred to the ancestors of green or land plants. Available experimental evidence shows that these anciently acquired genes are involved in some essential or plant-specific activities such as xylem formation, plant defence, nitrogen recycling as well as the biosynthesis of starch, polyamines, hormones and glutathione. These findings suggest that horizontal gene transfer had a critical role in the transition of plants from aquatic to terrestrial environments. On the basis of these findings, we propose a model of horizontal gene transfer mechanism in nonvascular and seedless vascular plants. PMID:23093189

  11. Widespread impact of horizontal gene transfer on plant colonization of land.

    PubMed

    Yue, Jipei; Hu, Xiangyang; Sun, Hang; Yang, Yongping; Huang, Jinling

    2012-01-01

    In complex multicellular eukaryotes such as animals and plants, horizontal gene transfer is commonly considered rare with very limited evolutionary significance. Here we show that horizontal gene transfer is a dynamic process occurring frequently in the early evolution of land plants. Our genome analyses of the moss Physcomitrella patens identified 57 families of nuclear genes that were acquired from prokaryotes, fungi or viruses. Many of these gene families were transferred to the ancestors of green or land plants. Available experimental evidence shows that these anciently acquired genes are involved in some essential or plant-specific activities such as xylem formation, plant defence, nitrogen recycling as well as the biosynthesis of starch, polyamines, hormones and glutathione. These findings suggest that horizontal gene transfer had a critical role in the transition of plants from aquatic to terrestrial environments. On the basis of these findings, we propose a model of horizontal gene transfer mechanism in nonvascular and seedless vascular plants.

  12. 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.

  13. Evolutionary implications of phylogenetic analyses of the gene transfer agent (GTA) of Rhodobacter capsulatus.

    PubMed

    Lang, Andrew S; Taylor, Terumi A; Beatty, J Thomas

    2002-11-01

    The gene transfer agent (GTA) of the a-proteobacterium Rhodobacter capsulatus is a cell-controlled genetic exchange vector. Genes that encode the GTA structure are clustered in a 15-kb region of the R. capsulatus chromosome, and some of these genes show sequence similarity to known bacteriophage head and tail genes. However, the production of GTA is controlled at the level of transcription by a cellular two-component signal transduction system. This paper describes homologues of both the GTA structural gene cluster and the GTA regulatory genes in the a-proteobacteria Rhodopseudomonas palustris, Rhodobacter sphaeroides, Caulobacter crescentus, Agrobacterium tumefaciens and Brucella melitensis. These sequences were used in a phylogenetic tree approach to examine the evolutionary relationships of selected GTA proteins to these homologues and (pro)phage proteins, which was compared to a 16S rRNA tree. The data indicate that a GTA-like element was present in a single progenitor of the extant species that contain both GTA structural cluster and regulatory gene homologues. The evolutionary relationships of GTA structural proteins to (pro)phage proteins indicated by the phylogenetic tree patterns suggest a predominantly vertical descent of GTA-like sequences in the a-proteobacteria and little past gene exchange with (pro)phages.

  14. Satellite services system analysis study: Propellant transfer system

    NASA Technical Reports Server (NTRS)

    1982-01-01

    General servicing requirements, a servicing mission concept and scenario, overall servicing needs, basic servicing equipment, and a general servicing mission configuration layout are addressed. Servicing needs, equipment concepts, system requirements equipment specifications, preliminary designs, and resource requirements for flight hardware for the propellant transfer system are also addressed.

  15. The Ensembl gene annotation system

    PubMed Central

    Aken, Bronwen L.; Ayling, Sarah; Barrell, Daniel; Clarke, Laura; Curwen, Valery; Fairley, Susan; Fernandez Banet, Julio; Billis, Konstantinos; García Girón, Carlos; Hourlier, Thibaut; Howe, Kevin; Kähäri, Andreas; Kokocinski, Felix; Martin, Fergal J.; Murphy, Daniel N.; Nag, Rishi; Ruffier, Magali; Schuster, Michael; Tang, Y. Amy; Vogel, Jan-Hinnerk; White, Simon; Zadissa, Amonida; Flicek, Paul

    2016-01-01

    The Ensembl gene annotation system has been used to annotate over 70 different vertebrate species across a wide range of genome projects. Furthermore, it generates the automatic alignment-based annotation for the human and mouse GENCODE gene sets. The system is based on the alignment of biological sequences, including cDNAs, proteins and RNA-seq reads, to the target genome in order to construct candidate transcript models. Careful assessment and filtering of these candidate transcripts ultimately leads to the final gene set, which is made available on the Ensembl website. Here, we describe the annotation process in detail. Database URL: http://www.ensembl.org/index.html PMID:27337980

  16. 5 CFR 843.210 - Transfers between retirement systems.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 5 Administrative Personnel 2 2012-01-01 2012-01-01 false Transfers between retirement systems. 843... REGULATIONS (CONTINUED) FEDERAL EMPLOYEES RETIREMENT SYSTEM-DEATH BENEFITS AND EMPLOYEE REFUNDS One-time Payments § 843.210 Transfers between retirement systems. Transfers of employees' contributions between...

  17. 5 CFR 843.210 - Transfers between retirement systems.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 5 Administrative Personnel 2 2014-01-01 2014-01-01 false Transfers between retirement systems. 843... REGULATIONS (CONTINUED) FEDERAL EMPLOYEES RETIREMENT SYSTEM-DEATH BENEFITS AND EMPLOYEE REFUNDS One-time Payments § 843.210 Transfers between retirement systems. Transfers of employees' contributions between...

  18. 5 CFR 843.210 - Transfers between retirement systems.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 5 Administrative Personnel 2 2013-01-01 2013-01-01 false Transfers between retirement systems. 843... REGULATIONS (CONTINUED) FEDERAL EMPLOYEES RETIREMENT SYSTEM-DEATH BENEFITS AND EMPLOYEE REFUNDS One-time Payments § 843.210 Transfers between retirement systems. Transfers of employees' contributions between...

  19. Recalibrating the BC Transfer System: Approved Final Report

    ERIC Educational Resources Information Center

    British Columbia Council on Admissions and Transfer, 2006

    2006-01-01

    In November 2005, the BC Council on Admissions and Transfer launched a consultation entitled Recalibrating the BC Transfer System with the institutional members of the BC Transfer System and other interested parties. This consultation was motivated in large part by significant changes in the BC post-secondary system over the last decade, and…

  20. 40 CFR 63.689 - Standards: Transfer systems.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 11 2013-07-01 2013-07-01 false Standards: Transfer systems. 63.689....689 Standards: Transfer systems. (a) The provisions of this section apply to the control of air emissions from transfer systems for which § 63.683(b)(1)(i) of this subpart references the use of...

  1. Proton transfer in some periodic molecular systems.

    PubMed

    Chojnacki, Henryk

    2007-07-01

    The electronic structure of representative hydrogen bonded systems: hydrogen cyanide, imidazole and malonic acid have been studied at the non-empirical level. The role of the dimensionality on the potential barrier for the proton transfer has been examined. It was shown that it depends on the crystal structure and only in some cases like hydrogen cyanide or imidazole the relevant crystals may be considered as one-dimensional. However, for more complicated crystallographic structures, e.g. malonic acid, the evaluated barrier is strongly dependent on the dimensionality taken into account in our calculations.

  2. Gene-transfer study approval awaits more data

    SciTech Connect

    Marwick, C.

    1988-11-18

    Approval of the gene-transfer study in cancer patients has been delayed. The proposal was recommended for approval by a National Institutes of Health (NIH) advisory committee, but has been put on hold by James B. Wyngaarden, MD, NIH director, pending submission in writing of further information. Some of this information, now forthcoming, had been withheld because data on preliminary studies had been submitted to peer-reviewed journals. The study involves placing the gene for neomycin-resistance to tumor-infiltrating lymphocytes as a marker. When these cells are injected into the patient, the presence of the marker should enable their fate to be studied over a prolonged period and an improved antitumor regimen could result. The use of tumor-infiltrating lymphocytes as immunotherapy has been studied for two years at the NIH's National Cancer Institute. The patients' tumors are removed and the tumor-infiltrating lymphocytes are cultivated to obtain several billion cells. These cells are then injected back into the patient. Early clinical experience has shown a substantial decease in tumor size in some patients, but not in all, an no one knows why.

  3. Graphene materials as 2D non-viral gene transfer vector platforms.

    PubMed

    Vincent, M; de Lázaro, I; Kostarelos, K

    2017-01-05

    Advances in genomics and gene therapy could offer solutions to many diseases that remain incurable today, however, one of the critical reasons halting clinical progress is due to the difficulty in designing efficient and safe delivery vectors for the appropriate genetic cargo. Safety and large-scale production concerns counter-balance the high gene transfer efficiency achieved with viral vectors, while non-viral strategies have yet to become sufficiently efficient. The extraordinary physicochemical, optical and photothermal properties of graphene-based materials (GBMs) could offer two-dimensional components for the design of nucleic acid carrier systems. We discuss here such properties and their implications for the optimization of gene delivery. While the design of such vectors is still in its infancy, we provide here an exhaustive and up-to-date analysis of the studies that have explored GBMs as gene transfer vectors, focusing on the functionalization strategies followed to improve vector performance and on the biological effects attained.Gene Therapy advance online publication, 5 January 2017; doi:10.1038/gt.2016.79.

  4. Sleeping Beauty-Mediated Drug Resistance Gene Transfer in Human Hematopoietic Progenitor Cells

    PubMed Central

    Hyland, Kendra A.; Olson, Erik R.; McIvor, R. Scott

    2015-01-01

    The Sleeping Beauty (SB) transposon system can insert sequences into mammalian chromosomes, supporting long-term expression of both reporter and therapeutic genes. Hematopoietic progenitor cells (HPCs) are an ideal therapeutic gene transfer target as they are used in therapy for a variety of hematologic and metabolic conditions. As successful SB-mediated gene transfer into human CD34+ HPCs has been reported by several laboratories, we sought to extend these studies to the introduction of a therapeutic gene conferring resistance to methotrexate (MTX), potentially providing a chemoprotective effect after engraftment. SB-mediated transposition of hematopoietic progenitors, using a transposon encoding an L22Y variant dihydrofolate reductase fused to green fluorescent protein, conferred resistance to methotrexate and dipyridamole, a nucleoside transport inhibitor that tightens MTX selection conditions, as assessed by in vitro hematopoietic colony formation. Transposition of individual transgenes was confirmed by sequence analysis of transposon–chromosome junctions recovered by linear amplification-mediated PCR. These studies demonstrate the potential of SB-mediated transposition of HPCs for expression of drug resistance genes for selective and chemoprotective applications. PMID:26176276

  5. Multiple Inter-Kingdom Horizontal Gene Transfers in the Evolution of the Phosphoenolpyruvate Carboxylase Gene Family

    PubMed Central

    Wang, Wen; Su, Bing

    2012-01-01

    Pepcase is a gene encoding phosphoenolpyruvate carboxylase that exists in bacteria, archaea and plants,playing an important role in plant metabolism and development. Most plants have two or more pepcase genes belonging to two gene sub-families, while only one gene exists in other organisms. Previous research categorized one plant pepcase gene as plant-type pepcase (PTPC) while the other as bacteria-type pepcase (BTPC) because of its similarity with the pepcase gene found in bacteria. Phylogenetic reconstruction showed that PTPC is the ancestral lineage of plant pepcase, and that all bacteria, protistpepcase and BTPC in plants are derived from a lineage of pepcase closely related with PTPC in algae. However, their phylogeny contradicts the species tree and traditional chronology of organism evolution. Because the diversification of bacteria occurred much earlier than the origin of plants, presumably all bacterialpepcase derived from the ancestral PTPC of algal plants after divergingfrom the ancestor of vascular plant PTPC. To solve this contradiction, we reconstructed the phylogeny of pepcase gene family. Our result showed that both PTPC and BTPC are derived from an ancestral lineage of gamma-proteobacteriapepcases, possibly via an ancient inter-kingdom horizontal gene transfer (HGT) from bacteria to the eukaryotic common ancestor of plants, protists and cellular slime mold. Our phylogenetic analysis also found 48other pepcase genes originated from inter-kingdom HGTs. These results imply that inter-kingdom HGTs played important roles in the evolution of the pepcase gene family and furthermore that HGTsare a more frequent evolutionary event than previouslythought. PMID:23251445

  6. Frequent, independent transfers of a catabolic gene from bacteria to contrasted filamentous eukaryotes

    PubMed Central

    Bruto, Maxime; Prigent-Combaret, Claire; Luis, Patricia; Moënne-Loccoz, Yvan; Muller, Daniel

    2014-01-01

    Even genetically distant prokaryotes can exchange genes between them, and these horizontal gene transfer events play a central role in adaptation and evolution. While this was long thought to be restricted to prokaryotes, certain eukaryotes have acquired genes of bacterial origin. However, gene acquisitions in eukaryotes are thought to be much less important in magnitude than in prokaryotes. Here, we describe the complex evolutionary history of a bacterial catabolic gene that has been transferred repeatedly from different bacterial phyla to stramenopiles and fungi. Indeed, phylogenomic analysis pointed to multiple acquisitions of the gene in these filamentous eukaryotes—as many as 15 different events for 65 microeukaryotes. Furthermore, once transferred, this gene acquired introns and was found expressed in mRNA databases for most recipients. Our results show that effective inter-domain transfers and subsequent adaptation of a prokaryotic gene in eukaryotic cells can happen at an unprecedented magnitude. PMID:24990676

  7. Frequent, independent transfers of a catabolic gene from bacteria to contrasted filamentous eukaryotes.

    PubMed

    Bruto, Maxime; Prigent-Combaret, Claire; Luis, Patricia; Moënne-Loccoz, Yvan; Muller, Daniel

    2014-08-22

    Even genetically distant prokaryotes can exchange genes between them, and these horizontal gene transfer events play a central role in adaptation and evolution. While this was long thought to be restricted to prokaryotes, certain eukaryotes have acquired genes of bacterial origin. However, gene acquisitions in eukaryotes are thought to be much less important in magnitude than in prokaryotes. Here, we describe the complex evolutionary history of a bacterial catabolic gene that has been transferred repeatedly from different bacterial phyla to stramenopiles and fungi. Indeed, phylogenomic analysis pointed to multiple acquisitions of the gene in these filamentous eukaryotes-as many as 15 different events for 65 microeukaryotes. Furthermore, once transferred, this gene acquired introns and was found expressed in mRNA databases for most recipients. Our results show that effective inter-domain transfers and subsequent adaptation of a prokaryotic gene in eukaryotic cells can happen at an unprecedented magnitude.

  8. Selective gene transfer to endometrial cancer cells by a polymer against matrix metalloproteinase 2 (MMP-2).

    PubMed

    Han, Joo Youn; Choi, Dong Soon; Kim, Changhoon; Joo, Hyun; Min, Churl K

    2008-04-01

    A novel cancer-cell-specific gene delivery vector with high transfection efficiency was designed and tested with an in vitro coculture consisting of the human endometrial adenocarcinoma cell line, HEC-1A cells, and normal endometrial stromal cells. For the cancer-cell targeting, polyethylenimine (PEI), a cationic polymer that can be easily combined with anionic DNA to form a particulate complex, polyplex, being capable of transferring a gene into a variety of cells, was covalently conjugated with antibodies against matrix metalloproteinase 2 (MMP-2), a typical surface-marker protein on cancer cells known for its close correlation with angiogenesis and invasion in many types of cancer, using the heterofunctional cross-linker, n-succinimidyl 3-(2-pyridyldithio)-propionamide. Biophysical properties and transfection efficiencies of anti-MMP-2-conjugated PEI were analyzed by means of dynamic light scattering, laser Doppler anemometry, and flow cytometry. Our results reveal that (1) the PEI-anti-MMP-2 antibody conjugate maintains physical parameters, including sizes and surface charges, which appear to be favorable for gene transfer and (2) when the pEGFP-N3 plasmid complexes of the PEI-anti-MMP-2 antibody conjugate are applied to the coculture consisting of HEC-1A cells and human stromal cells, a high level of green fluorescent protein expression occurs in HEC-1A cells over stromal cells, suggesting a specific gene transfer targeting cancer cells. Therefore, targeting invading cancer cells with the PEI-anti-MMP-2 antibody conjugate could be promising in endometrial cancer treatment, and this gene delivery system deserves further optimization in the context of targeted therapeutic gene delivery.

  9. Center for fetal monkey gene transfer for heart, lung, and blood diseases: an NHLBI resource for the gene therapy community.

    PubMed

    Tarantal, Alice F; Skarlatos, Sonia I

    2012-11-01

    The goals of the National Heart, Lung, and Blood Institute (NHLBI) Center for Fetal Monkey Gene Transfer for Heart, Lung, and Blood Diseases are to conduct gene transfer studies in monkeys to evaluate safety and efficiency; and to provide NHLBI-supported investigators with expertise, resources, and services to actively pursue gene transfer approaches in monkeys in their research programs. NHLBI-supported projects span investigators throughout the United States and have addressed novel approaches to gene delivery; "proof-of-principle"; assessed whether findings in small-animal models could be demonstrated in a primate species; or were conducted to enable new grant or IND submissions. The Center for Fetal Monkey Gene Transfer for Heart, Lung, and Blood Diseases successfully aids the gene therapy community in addressing regulatory barriers, and serves as an effective vehicle for advancing the field.

  10. [Gene transfer agent--a novel and widespread occurrence mechanism of gene exchange in ocean-a review].

    PubMed

    Cai, Haiyuan

    2012-01-01

    Gene Transfer Agent (GTA) particles are released by bacteria and resemble small, tailed bacteriophages. GTA particles contain small, random pieces of host DNA rather than GTA structural genes or a phage genome. Gene transfer mediated by GTA is efficient and species specific based on knowledge of currently best studied GTAs produced by 4 anaerobes. Genome sequencing projects have revealed a remarkable distribution of GTA gene clusters in the genomes of marine bacterioplankton, implying GTA may be an important mechanism for horizontal gene transfer in ocean. On basis of characterization of the 4 best studied GTAs, this review described GTAs released by numerically dominant marine bacteria, discussed their properties that were important for horizontal gene transfer in ocean, and gave future perspectives to advance GTA research.

  11. The elucidation of gene transferring mechanism by ultrasound-responsive unmodified and mannose-modified lipoplexes.

    PubMed

    Un, Keita; Kawakami, Shigeru; Yoshida, Mitsuru; Higuchi, Yuriko; Suzuki, Ryo; Maruyama, Kazuo; Yamashita, Fumiyoshi; Hashida, Mitsuru

    2011-07-01

    The development of gene transfection methods enhancing the level of gene expression under simple and low-toxic condition is required for gene therapy in clinical. Our group has developed the ultrasound (US)-mediated gene transfection method using Man-PEG(2000) bubble lipoplexes, which are US-responsive and mannose-modified gene carriers, and succeeded in obtaining the enhanced gene expression in mannose receptor-expressing cells selectively by the gene transfer using Man-PEG(2000) bubble lipoplexes with US exposure in vitro and in vivo. Here, we investigated pDNA transferring mechanism followed by US exposure to unmodified and Man-PEG(2000) bubble lipoplexes, in particular, focused on US exposure timing. Following investigation of intracellular transferring characteristics, a large amount of pDNA was transferred into the cytoplasm followed by US-mediated destruction of bubble lipoplexes in the gene transfer using both bubble lipoplexes with US exposure. Moreover, the effective gene expression was obtained without TNF-α production when US was exposed until 5 min after the addition of bubble lipoplexes. These findings suggest that the gene transfer using unmodified and Man-PEG(2000) bubble lipoplexes with US exposure enables to transfer pDNA into the cytoplasm, and optimized US exposure timing is important to achieve the high level of gene expression and the low level of pro-inflammatory cytokine production.

  12. Transfer zones in listric normal fault systems

    NASA Astrophysics Data System (ADS)

    Bose, Shamik

    Listric normal faults are common in passive margin settings where sedimentary units are detached above weaker lithological units, such as evaporites or are driven by basal structural and stratigraphic discontinuities. The geometries and styles of faulting vary with the types of detachment and form landward and basinward dipping fault systems. Complex transfer zones therefore develop along the terminations of adjacent faults where deformation is accommodated by secondary faults, often below seismic resolution. The rollover geometry and secondary faults within the hanging wall of the major faults also vary with the styles of faulting and contribute to the complexity of the transfer zones. This study tries to understand the controlling factors for the formation of the different styles of listric normal faults and the different transfer zones formed within them, by using analog clay experimental models. Detailed analyses with respect to fault orientation, density and connectivity have been performed on the experiments in order to gather insights on the structural controls and the resulting geometries. A new high resolution 3D laser scanning technology has been introduced to scan the surfaces of the clay experiments for accurate measurements and 3D visualizations. Numerous examples from the Gulf of Mexico have been included to demonstrate and geometrically compare the observations in experiments and real structures. A salt cored convergent transfer zone from the South Timbalier Block 54, offshore Louisiana has been analyzed in detail to understand the evolutionary history of the region, which helps in deciphering the kinematic growth of similar structures in the Gulf of Mexico. The dissertation is divided into three chapters, written in a journal article format, that deal with three different aspects in understanding the listric normal fault systems and the transfer zones so formed. The first chapter involves clay experimental models to understand the fault patterns in

  13. Targeting adipose tissue via systemic gene therapy.

    PubMed

    O'Neill, S M; Hinkle, C; Chen, S-J; Sandhu, A; Hovhannisyan, R; Stephan, S; Lagor, W R; Ahima, R S; Johnston, J C; Reilly, M P

    2014-07-01

    Adipose tissue has a critical role in energy and metabolic homeostasis, but it is challenging to adapt techniques to modulate adipose function in vivo. Here we develop an in vivo, systemic method of gene transfer specifically targeting adipose tissue using adeno-associated virus (AAV) vectors. We constructed AAV vectors containing cytomegalovirus promoter-regulated reporter genes, intravenously injected adult mice with vectors using multiple AAV serotypes, and determined that AAV2/8 best targeted adipose tissue. Altering vectors to contain adiponectin promoter/enhancer elements and liver-specific microRNA-122 target sites restricted reporter gene expression to adipose tissue. As proof of efficacy, the leptin gene was incorporated into the adipose-targeted expression vector, package into AAV2/8 and administered intravenously to 9- to 10-week-old ob/ob mice. Phenotypic changes were measured over an 8-week period. Leptin mRNA and protein were expressed in adipose and leptin protein was secreted into plasma. Mice responded with reversal of weight gain, decreased hyperinsulinemia and improved glucose tolerance. AAV2/8-mediated systemic delivery of an adipose-targeted expression vector can replace a gene lacking in adipose tissue and correct a mouse model of human disease, demonstrating experimental application and therapeutic potential in disorders of adipose.

  14. Horizontal gene transfer of chlamydial-like tRNA genes into early vascular plant mitochondria.

    PubMed

    Knie, Nils; Polsakiewicz, Monika; Knoop, Volker

    2015-03-01

    Mitochondrial genomes of lycophytes are surprisingly diverse, including strikingly different transfer RNA (tRNA) gene complements: No mitochondrial tRNA genes are present in the spikemoss Selaginella moellendorffii, whereas 26 tRNAs are encoded in the chondrome of the clubmoss Huperzia squarrosa. Reinvestigating the latter we found that trnL(gag) and trnS(gga) had never before been identified in any other land plant mitochondrial DNA. Sensitive sequence comparisons showed these two tRNAs as well as trnN(guu) and trnS(gcu) to be very similar to their respective counterparts in chlamydial bacteria. We identified homologs of these chlamydial-type tRNAs also in other lycophyte, fern, and gymnosperm DNAs, suggesting horizontal gene transfer (HGT) into mitochondria in the early vascular plant stem lineages. These findings extend plant mitochondrial HGT to affect individual tRNA genes, to include bacterial donors, and suggest that Chlamydiae on top of their recently proposed key role in primary chloroplast establishment may also have participated in early tracheophyte genome evolution.

  15. 46 CFR 153.430 - Heat transfer systems; general.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 46 Shipping 5 2013-10-01 2013-10-01 false Heat transfer systems; general. 153.430 Section 153.430... Temperature Control Systems § 153.430 Heat transfer systems; general. Each cargo cooling system required by... separated from all other cooling and heating systems; and (c) Allow manual regulation of the system's...

  16. 46 CFR 153.430 - Heat transfer systems; general.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... 46 Shipping 5 2012-10-01 2012-10-01 false Heat transfer systems; general. 153.430 Section 153.430... Temperature Control Systems § 153.430 Heat transfer systems; general. Each cargo cooling system required by... separated from all other cooling and heating systems; and (c) Allow manual regulation of the system's...

  17. 46 CFR 153.430 - Heat transfer systems; general.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 46 Shipping 5 2014-10-01 2014-10-01 false Heat transfer systems; general. 153.430 Section 153.430... Temperature Control Systems § 153.430 Heat transfer systems; general. Each cargo cooling system required by... separated from all other cooling and heating systems; and (c) Allow manual regulation of the system's...

  18. 46 CFR 153.430 - Heat transfer systems; general.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 46 Shipping 5 2011-10-01 2011-10-01 false Heat transfer systems; general. 153.430 Section 153.430... Temperature Control Systems § 153.430 Heat transfer systems; general. Each cargo cooling system required by... separated from all other cooling and heating systems; and (c) Allow manual regulation of the system's...

  19. 46 CFR 153.430 - Heat transfer systems; general.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 46 Shipping 5 2010-10-01 2010-10-01 false Heat transfer systems; general. 153.430 Section 153.430... Temperature Control Systems § 153.430 Heat transfer systems; general. Each cargo cooling system required by... separated from all other cooling and heating systems; and (c) Allow manual regulation of the system's...

  20. Horizontal gene transfer and gene dosage drives adaptation to wood colonization in a tree pathogen.

    PubMed

    Dhillon, Braham; Feau, Nicolas; Aerts, Andrea L; Beauseigle, Stéphanie; Bernier, Louis; Copeland, Alex; Foster, Adam; Gill, Navdeep; Henrissat, Bernard; Herath, Padmini; LaButti, Kurt M; Levasseur, Anthony; Lindquist, Erika A; Majoor, Eline; Ohm, Robin A; Pangilinan, Jasmyn L; Pribowo, Amadeus; Saddler, John N; Sakalidis, Monique L; de Vries, Ronald P; Grigoriev, Igor V; Goodwin, Stephen B; Tanguay, Philippe; Hamelin, Richard C

    2015-03-17

    Some of the most damaging tree pathogens can attack woody stems, causing lesions (cankers) that may be lethal. To identify the genomic determinants of wood colonization leading to canker formation, we sequenced the genomes of the poplar canker pathogen, Mycosphaerella populorum, and the closely related poplar leaf pathogen, M. populicola. A secondary metabolite cluster unique to M. populorum is fully activated following induction by poplar wood and leaves. In addition, genes encoding hemicellulose-degrading enzymes, peptidases, and metabolite transporters were more abundant and were up-regulated in M. populorum growing on poplar wood-chip medium compared with M. populicola. The secondary gene cluster and several of the carbohydrate degradation genes have the signature of horizontal transfer from ascomycete fungi associated with wood decay and from prokaryotes. Acquisition and maintenance of the gene battery necessary for growth in woody tissues and gene dosage resulting in gene expression reconfiguration appear to be responsible for the adaptation of M. populorum to infect, colonize, and cause mortality on poplar woody stems.

  1. Horizontal gene transfer and gene dosage drives adaptation to wood colonization in a tree pathogen

    PubMed Central

    Dhillon, Braham; Feau, Nicolas; Aerts, Andrea L.; Beauseigle, Stéphanie; Bernier, Louis; Copeland, Alex; Foster, Adam; Gill, Navdeep; Henrissat, Bernard; Herath, Padmini; LaButti, Kurt M.; Levasseur, Anthony; Lindquist, Erika A.; Majoor, Eline; Ohm, Robin A.; Pangilinan, Jasmyn L.; Pribowo, Amadeus; Saddler, John N.; Sakalidis, Monique L.; de Vries, Ronald P.; Grigoriev, Igor V.; Goodwin, Stephen B.; Tanguay, Philippe; Hamelin, Richard C.

    2015-01-01

    Some of the most damaging tree pathogens can attack woody stems, causing lesions (cankers) that may be lethal. To identify the genomic determinants of wood colonization leading to canker formation, we sequenced the genomes of the poplar canker pathogen, Mycosphaerella populorum, and the closely related poplar leaf pathogen, M. populicola. A secondary metabolite cluster unique to M. populorum is fully activated following induction by poplar wood and leaves. In addition, genes encoding hemicellulose-degrading enzymes, peptidases, and metabolite transporters were more abundant and were up-regulated in M. populorum growing on poplar wood-chip medium compared with M. populicola. The secondary gene cluster and several of the carbohydrate degradation genes have the signature of horizontal transfer from ascomycete fungi associated with wood decay and from prokaryotes. Acquisition and maintenance of the gene battery necessary for growth in woody tissues and gene dosage resulting in gene expression reconfiguration appear to be responsible for the adaptation of M. populorum to infect, colonize, and cause mortality on poplar woody stems. PMID:25733908

  2. Transduction-Like Gene Transfer in the Methanogen Methanococcus voltae

    PubMed Central

    Bertani, Giuseppe

    1999-01-01

    Strain PS of Methanococcus voltae (a methanogenic, anaerobic archaebacterium) was shown to generate spontaneously 4.4-kbp chromosomal DNA fragments that are fully protected from DNase and that, upon contact with a cell, transform it genetically. This activity, here called VTA (voltae transfer agent), affects all markers tested: three different auxotrophies (histidine, purine, and cobalamin) and resistance to BES (2-bromoethanesulfonate, an inhibitor of methanogenesis). VTA was most effectively prepared by culture filtration. This process disrupted a fraction of the M. voltae cells (which have only an S-layer covering their cytoplasmic membrane). VTA was rapidly inactivated upon storage. VTA particles were present in cultures at concentrations of approximately two per cell. Gene transfer activity varied from a minimum of 2 × 10−5 (BES resistance) to a maximum of 10−3 (histidine independence) per donor cell. Very little VTA was found free in culture supernatants. The phenomenon is functionally similar to generalized transduction, but there is no evidence, for the time being, of intrinsically viral (i.e., containing a complete viral genome) particles. Consideration of VTA DNA size makes the existence of such viral particles unlikely. If they exist, they must be relatively few in number;perhaps they differ from VTA particles in size and other properties and thus escaped detection. Digestion of VTA DNA with the AluI restriction enzyme suggests that it is a random sample of the bacterial DNA, except for a 0.9-kbp sequence which is amplified relative to the rest of the bacterial chromosome. A VTA-sized DNA fraction was demonstrated in a few other isolates of M. voltae. PMID:10321998

  3. Parvalbumin gene transfer impairs skeletal muscle contractility in old mice.

    PubMed

    Murphy, Kate T; Ham, Daniel J; Church, Jarrod E; Naim, Timur; Trieu, Jennifer; Williams, David A; Lynch, Gordon S

    2012-08-01

    Sarcopenia is the progressive age-related loss of skeletal muscle mass associated with functional impairments that reduce mobility and quality of life. Overt muscle wasting with sarcopenia is usually preceded by a slowing of the rate of relaxation and a reduction in maximum force production. Parvalbumin (PV) is a cytosolic Ca(2+) buffer thought to facilitate relaxation in muscle. We tested the hypothesis that restoration of PV levels in muscles of old mice would increase the magnitude and hasten relaxation of submaximal and maximal force responses. The tibialis anterior (TA) muscles of young (6 month), adult (13 month), and old (26 month) C57BL/6 mice received electroporation-assisted gene transfer of plasmid encoding PV or empty plasmid (pcDNA3.1). Contractile properties of TA muscles were assessed in situ 14 days after transfer. In old mice, muscles with increased PV expression had a 40% slower rate of tetanic force development (p<0.01), and maximum twitch and tetanic force were 22% and 16% lower than control values, respectively (p<0.05). Muscles with increased PV expression from old mice had an 18% lower maximum specific (normalized) force than controls, and absolute force was `26% lower at higher stimulation frequencies (150-300 Hz, p<0.05). In contrast, there was no effect of increased PV expression on TA muscle contractile properties in young and adult mice. The impairments in skeletal muscle function in old mice argue against PV overexpression as a therapeutic strategy for ameliorating aspects of contractile dysfunction with sarcopenia and help clarify directions for therapeutic interventions for age-related changes in skeletal muscle structure and function.

  4. Novel "Superspreader" Bacteriophages Promote Horizontal Gene Transfer by Transformation.

    PubMed

    Keen, Eric C; Bliskovsky, Valery V; Malagon, Francisco; Baker, James D; Prince, Jeffrey S; Klaus, James S; Adhya, Sankar L

    2017-01-17

    Bacteriophages infect an estimated 10(23) to 10(25) bacterial cells each second, many of which carry physiologically relevant plasmids (e.g., those encoding antibiotic resistance). However, even though phage-plasmid interactions occur on a massive scale and have potentially significant evolutionary, ecological, and biomedical implications, plasmid fate upon phage infection and lysis has not been investigated to date. Here we show that a subset of the natural lytic phage population, which we dub "superspreaders," releases substantial amounts of intact, transformable plasmid DNA upon lysis, thereby promoting horizontal gene transfer by transformation. Two novel Escherichia coli phage superspreaders, SUSP1 and SUSP2, liberated four evolutionarily distinct plasmids with equal efficiency, including two close relatives of prominent antibiotic resistance vectors in natural environments. SUSP2 also mediated the extensive lateral transfer of antibiotic resistance in unbiased communities of soil bacteria from Maryland and Wyoming. Furthermore, the addition of SUSP2 to cocultures of kanamycin-resistant E. coli and kanamycin-sensitive Bacillus sp. bacteria resulted in roughly 1,000-fold more kanamycin-resistant Bacillus sp. bacteria than arose in phage-free controls. Unlike many other lytic phages, neither SUSP1 nor SUSP2 encodes homologs to known hydrolytic endonucleases, suggesting a simple potential mechanism underlying the superspreading phenotype. Consistent with this model, the deletion of endonuclease IV and the nucleoid-disrupting protein ndd from coliphage T4, a phage known to extensively degrade chromosomal DNA, significantly increased its ability to promote plasmid transformation. Taken together, our results suggest that phage superspreaders may play key roles in microbial evolution and ecology but should be avoided in phage therapy and other medical applications.

  5. Transduction-like gene transfer in the methanogen Methanococcus voltae

    NASA Technical Reports Server (NTRS)

    Bertani, G.

    1999-01-01

    Strain PS of Methanococcus voltae (a methanogenic, anaerobic archaebacterium) was shown to generate spontaneously 4.4-kbp chromosomal DNA fragments that are fully protected from DNase and that, upon contact with a cell, transform it genetically. This activity, here called VTA (voltae transfer agent), affects all markers tested: three different auxotrophies (histidine, purine, and cobalamin) and resistance to BES (2-bromoethanesulfonate, an inhibitor of methanogenesis). VTA was most effectively prepared by culture filtration. This process disrupted a fraction of the M. voltae cells (which have only an S-layer covering their cytoplasmic membrane). VTA was rapidly inactivated upon storage. VTA particles were present in cultures at concentrations of approximately two per cell. Gene transfer activity varied from a minimum of 2 x 10(-5) (BES resistance) to a maximum of 10(-3) (histidine independence) per donor cell. Very little VTA was found free in culture supernatants. The phenomenon is functionally similar to generalized transduction, but there is no evidence, for the time being, of intrinsically viral (i.e., containing a complete viral genome) particles. Consideration of VTA DNA size makes the existence of such viral particles unlikely. If they exist, they must be relatively few in number;perhaps they differ from VTA particles in size and other properties and thus escaped detection. Digestion of VTA DNA with the AluI restriction enzyme suggests that it is a random sample of the bacterial DNA, except for a 0.9-kbp sequence which is amplified relative to the rest of the bacterial chromosome. A VTA-sized DNA fraction was demonstrated in a few other isolates of M. voltae.

  6. 5 CFR 831.2010 - Transfers between retirement systems.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 5 Administrative Personnel 2 2012-01-01 2012-01-01 false Transfers between retirement systems. 831... REGULATIONS (CONTINUED) RETIREMENT Payment of Lump Sums § 831.2010 Transfers between retirement systems. Transfers of employees' contributions between the Civil Service Retirement and Disability Fund and...

  7. 5 CFR 831.2010 - Transfers between retirement systems.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 5 Administrative Personnel 2 2013-01-01 2013-01-01 false Transfers between retirement systems. 831... REGULATIONS (CONTINUED) RETIREMENT Payment of Lump Sums § 831.2010 Transfers between retirement systems. Transfers of employees' contributions between the Civil Service Retirement and Disability Fund and...

  8. 5 CFR 831.2010 - Transfers between retirement systems.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 5 Administrative Personnel 2 2011-01-01 2011-01-01 false Transfers between retirement systems. 831... REGULATIONS (CONTINUED) RETIREMENT Payment of Lump Sums § 831.2010 Transfers between retirement systems. Transfers of employees' contributions between the Civil Service Retirement and Disability Fund and...

  9. 5 CFR 831.2010 - Transfers between retirement systems.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 5 Administrative Personnel 2 2014-01-01 2014-01-01 false Transfers between retirement systems. 831... REGULATIONS (CONTINUED) RETIREMENT Payment of Lump Sums § 831.2010 Transfers between retirement systems. Transfers of employees' contributions between the Civil Service Retirement and Disability Fund and...

  10. 5 CFR 831.2010 - Transfers between retirement systems.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 5 Administrative Personnel 2 2010-01-01 2010-01-01 false Transfers between retirement systems. 831... REGULATIONS (CONTINUED) RETIREMENT Payment of Lump Sums § 831.2010 Transfers between retirement systems. Transfers of employees' contributions between the Civil Service Retirement and Disability Fund and...

  11. Multimodality Imaging of Gene Transfer with a Receptor-Based Reporter Gene

    PubMed Central

    Chen, Ron; Parry, Jesse J.; Akers, Walter J.; Berezin, Mikhail Y.; El Naqa, Issam M.; Achilefu, Samuel; Edwards, W. Barry; Rogers, Buck E.

    2010-01-01

    Gene therapy trials have traditionally used tumor and tissue biopsies for assessing the efficacy of gene transfer. Non-invasive imaging techniques offer a distinct advantage over tissue biopsies in that the magnitude and duration of gene transfer can be monitored repeatedly. Human somatostatin receptor subtype 2 (SSTR2) has been used for the nuclear imaging of gene transfer. To extend this concept, we have developed a somatostatin receptor–enhanced green fluorescent protein fusion construct (SSTR2-EGFP) for nuclear and fluorescent multimodality imaging. Methods An adenovirus containing SSTR2-EGFP (AdSSTR2-EGFP) was constructed and evaluated in vitro and in vivo. SCC-9 human squamous cell carcinoma cells were infected with AdEGFP, AdSSTR2, or AdSSTR2-EGFP for in vitro evaluation by saturation binding, internalization, and fluorescence spectroscopy assays. In vivo biodistribution and nano-SPECT imaging studies were conducted with mice bearing SCC-9 tumor xenografts directly injected with AdSSTR2-EGFP or AdSSTR2 to determine the tumor localization of 111In-diethylenetriaminepentaacetic acid (DTPA)-Tyr3-octreotate. Fluorescence imaging was conducted in vivo with mice receiving intratumoral injections of AdSSTR2, AdSSTR2-EGFP, or AdEGFP as well as ex vivo with tissues extracted from mice. Results The similarity between AdSSTR2-EGFP and wild-type AdSSTR2 was demonstrated in vitro by the saturation binding and internalization assays, and the fluorescence emission spectra of cells infected with AdSSTR2-EGFP was almost identical to the spectra of cells infected with wild-type AdEGFP. Biodistribution studies demonstrated that the tumor uptake of 111In-DTPA-Tyr3-octreotate was not significantly different (P > 0.05) when tumors (n = 5) were injected with AdSSTR2 or AdSSTR2-EGFP but was significantly greater than the uptake in control tumors. Fluorescence was observed in tumors injected with AdSSTR2-EGFP and AdEGFP in vivo and ex vivo but not in tumors injected with AdSSTR2

  12. Microbial Evolution Is in the Cards: Horizontal Gene Transfer in the Classroom

    ERIC Educational Resources Information Center

    Kagle, Jeanne; Hay, Anthony G.

    2007-01-01

    Horizontal gene transfer, the exchange of genetic material between bacteria, is a potentially important factor in the degradation of synthetic compounds introduced to the environment and in the acquisition of other characteristics including antibiotic resistance. This game-based activity illustrates the role of horizontal gene transfer in the…

  13. Generation of CRISPR/Cas9-mediated gene-targeted pigs via somatic cell nuclear transfer.

    PubMed

    Zhou, Xiaoqing; Xin, Jige; Fan, Nana; Zou, Qingjian; Huang, Jiao; Ouyang, Zhen; Zhao, Yu; Zhao, Bentian; Liu, Zhaoming; Lai, Sisi; Yi, Xiaoling; Guo, Lin; Esteban, Miguel A; Zeng, Yangzhi; Yang, Huaqiang; Lai, Liangxue

    2015-03-01

    The domestic pig has been widely used as an important large animal model. Precise and efficient genetic modification in pig provides a great promise in biomedical research. Recently, clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated (Cas) system has been successfully used to produce many gene-targeted animals. However, these animals have been generated by co-injection of Cas9 mRNA and single-guide RNA (sgRNA) into one-cell stage embryos, which mostly resulted in mosaicism of the modification. One or two rounds of further breeding should be performed to obtain homozygotes with identical genotype and phenotype. To address this issue, gene-targeted somatic cells can be used as donor for somatic cell nuclear transfer (SCNT) to produce gene-targeted animals with single and identical mutations. In this study, we applied Cas9/sgRNAs to effectively direct gene editing in porcine fetal fibroblasts and then mutant cell colonies were used as donor to generate homozygous gene-targeted pigs through single round of SCNT. As a result, we successfully obtained 15 tyrosinase (TYR) biallelic mutant pigs and 20 PARK2 and PINK1 double-gene knockout (KO) pigs. They were all homozygous and no off-target mutagenesis was detected by comprehensive analysis. TYR (-/-) pigs showed typical albinism and the expression of parkin and PINK1 were depleted in PARK2 (-/-)/PINK1 (-/-) pigs. The results demonstrated that single- or double-gene targeted pigs can be effectively achieved by using the CRISPR/Cas9 system combined with SCNT without mosaic mutation and detectable off-target effects. This gene-editing system provides an efficient, rapid, and less costly manner to generate genetically modified pigs or other large animals.

  14. Targeted gene transfer of different genes to presynaptic and postsynaptic neocortical neurons connected by a glutamatergic synapse.

    PubMed

    Zhang, Guo-rong; Zhao, Hua; Cao, Haiyan; Li, Xu; Geller, Alfred I

    2012-09-14

    Genetic approaches to analyzing neuronal circuits and learning would benefit from a technology to first deliver a specific gene into presynaptic neurons, and then deliver a different gene into an identified subset of their postsynaptic neurons, connected by a specific synapse type. Here, we describe targeted gene transfer across a neocortical glutamatergic synapse, using as the model the projection from rat postrhinal to perirhinal cortex. The first gene transfer, into the presynaptic neurons in postrhinal cortex, used a virus vector and standard gene transfer procedures. The vector expresses an artificial peptide neurotransmitter containing a dense core vesicle targeting domain, a NMDA NR1 subunit binding domain (from a monoclonal antibody), and the His tag. Upon release, this peptide neurotransmitter binds to NMDA receptors on the postsynaptic neurons. Antibody-mediated targeted gene transfer to these postsynaptic neurons in perirhinal cortex used a His tag antibody, as the peptide neurotransmitter contains the His tag. Confocal microscopy showed that with untargeted gene transfer, ~3% of the transduced presynaptic axons were proximal to a transduced postsynaptic dendrite. In contrast, with targeted gene transfer, ≥ 20% of the presynaptic axons were proximal to a transduced postsynaptic dendrite. Targeting across other types of synapses might be obtained by modifying the artificial peptide neurotransmitter to contain a binding domain for a different neurotransmitter receptor. This technology may benefit elucidating how specific neurons and subcircuits contribute to circuit physiology, behavior, and learning.

  15. Persistent gene expression in mouse nasal epithelia following feline immunodeficiency virus-based vector gene transfer.

    PubMed

    Sinn, Patrick L; Burnight, Erin R; Hickey, Melissa A; Blissard, Gary W; McCray, Paul B

    2005-10-01

    Gene transfer development for treatment or prevention of cystic fibrosis lung disease has been limited by the inability of vectors to efficiently and persistently transduce airway epithelia. Influenza A is an enveloped virus with natural lung tropism; however, pseudotyping feline immunodeficiency virus (FIV)-based lentiviral vector with the hemagglutinin envelope protein proved unsuccessful. Conversely, pseudotyping FIV with the envelope protein from influenza D (Thogoto virus GP75) resulted in titers of 10(6) transducing units (TU)/ml and conferred apical entry into well-differentiated human airway epithelial cells. Baculovirus GP64 envelope glycoproteins share sequence identity with influenza D GP75 envelope glycoproteins. Pseudotyping FIV with GP64 from three species of baculovirus resulted in titers of 10(7) to 10(9) TU/ml. Of note, GP64 from Autographa californica multicapsid nucleopolyhedrovirus resulted in high-titer FIV preparations (approximately 10(9) TU/ml) and conferred apical entry into polarized primary cultures of human airway epithelia. Using a luciferase reporter gene and bioluminescence imaging, we observed persistent gene expression from in vivo gene transfer in the mouse nose with A. californica GP64-pseudotyped FIV (AcGP64-FIV). Longitudinal bioluminescence analysis documented persistent expression in nasal epithelia for approximately 1 year without significant decline. According to histological analysis using a LacZ reporter gene, olfactory and respiratory epithelial cells were transduced. In addition, methylcellulose-formulated AcGP64-FIV transduced mouse nasal epithelia with much greater efficiency than similarly formulated vesicular stomatitis virus glycoprotein-pseudotyped FIV. These data suggest that AcGP64-FIV efficiently transduces and persistently expresses a transgene in nasal epithelia in the absence of agents that disrupt the cellular tight junction integrity.

  16. Horizontal gene transfer events reshape the global landscape of arm race between viruses and homo sapiens

    PubMed Central

    Chen, Dong-Sheng; Wu, Yi-Quan; Zhang, Wei; Jiang, San-Jie; Chen, Shan-Ze

    2016-01-01

    Horizontal gene transfer (HGT) drives the evolution of recipient organism particularly if it provides a novel function which enhances the fitness or its adaption to the environment. Virus-host co-evolution is attractive for studying co-evolutionary processes, since viruses strictly replicate inside of the host cells and thus their evolution is inexorably tangled with host biology. HGT, as a mechanism of co-evolution between human and viruses, has been widely documented, however, the roles HGT play during the interaction between human and viruses are still in their infancy. In this study, we performed a comprehensive analysis on the genes horizontally transferred between viruses and their corresponding human hosts. Our study suggests that the HGT genes in human are predominantly enriched in immune related GO terms while viral HGT genes are tend to be encoded by viruses which promote the invasion of immune system of hosts. Based on our results, it gives us a hint about the evolution trajectory of HGT events. Overall, our study suggests that the HGT between human and viruses are highly relevant to immune interaction and probably reshaped the arm race between hosts and viruses. PMID:27270140

  17. Thixotropic solutions enhance viral-mediated gene transfer to airway epithelia.

    PubMed

    Seiler, Michael P; Luner, Paul; Moninger, Thomas O; Karp, Philip H; Keshavjee, Shaf; Zabner, Joseph

    2002-08-01

    Adenovirus-mediated gene transfer to airway epithelia is inefficient in part because its receptor is absent on the apical surface of the airways. Targeting adenovirus to other receptors, increasing the viral concentration, and even prolonging the incubation time with adenovirus vectors can partially overcome the lack of receptors and facilitate gene transfer. Unfortunately, mucociliary clearance would prevent prolonged incubation time in vivo. Thixotropic solutions (TS) are gels that upon a vigorous shearing force reversibly become liquid. We hypothesized that formulating recombinant adenoviruses in TS would decrease virus clearance and thus enhance gene transfer to the airway epithelia. We found that clearance of virus-sized fluorescent beads by human airway epithelia in vitro and by monkey trachea in vivo were markedly decreased when the beads were formulated in TS compared with phosphate-buffered saline (PBS). Adenovirus formulated in TS significantly increased adenovirus-mediated gene transfer of a reporter gene in human airway epithelia in vitro and in murine airway epithelia in vivo. Furthermore, an adenovirus encoding the cystic fibrosis transmembrane regulator (CFTR) gene (AdCFTR) formulated in TS was more efficient in correcting the chloride transport defect in cystic fibrosis airway epithelia than AdCFTR formulated in PBS. These data indicate a novel strategy to augment the efficiency of gene transfer to the airways that may be applicable to a number of different gene transfer vectors and could be of value in gene transfer to cystic fibrosis (CF) airway epithelia in vivo.

  18. GENE TRANSFER FOR THE TREATMENT OF NEOVASCULAR OCULAR DISEASE (AN AMERICAN OPHTHALMOLOGICAL SOCIETY THESIS)

    PubMed Central

    Stout, John Timothy

    2006-01-01

    Purpose As vasoproliferative diseases account for a substantial fraction of worldwide blindness and share the activation of the angiogenic pathway as a common etiology, the expression of antiangiogenic proteins offers a promising means of treatment. This study was designed to develop viral vectors, harboring angiostatic genes, for the study and treatment of experimental proliferative ocular disease. A variety of methods (in vitro, ex vivo tissue, and in vivo) were employed to model the process of proliferation and test the effectiveness of these reagents. Methods Antiangiogenic genes included single genes as well as hybrid genes that fused the active elements of different genes. Genes studied included the soluble vascular endothelial growth factor receptor (sKDR), soluble neuropilin (sNRP-1), tissue inhibitor of metalloproteinase-1 (TIMP-1), plasminogen gene fragments (Kringle 1-3, 1-4, and 1-5), and soluble receptors for advanced glycosylation end products (sRAGE) genes, as well as the Endo:Ang, MIG:IP10, and Endo:Kringle5 fusion genes. All genes were cloned into a lentiviral vector system and were used to produce replication deficient lentiviral particles. These viral particles were used to transduce a variety of ocular cells and tissues to test viral transfer efficiency and transgene expression. In vivo systems were employed to explore the potential of these genes as antiangiogenic agents in models of corneal and retinal neovascular disease. Results Recombinant lentiviral particles, capable of transducing cell lines germane to eye disease (ocular endothelial, epithelial, and fibroblast cells), were successfully produced. These vectors were demonstrated to be effective in long-term transformation of cells and tissues. In vivo experiments confirmed that at least three different potentially angiostatic genes were successful in aborting the angiogenic process in the ocular models tested. Conclusions Lentiviral vectors are a viable means to deliver angiostatic genes

  19. Horizontal gene transfer in the acquisition of novel traits by metazoans

    PubMed Central

    Boto, Luis

    2014-01-01

    Horizontal gene transfer is accepted as an important evolutionary force modulating the evolution of prokaryote genomes. However, it is thought that horizontal gene transfer plays only a minor role in metazoan evolution. In this paper, I critically review the rising evidence on horizontally transferred genes and on the acquisition of novel traits in metazoans. In particular, I discuss suspected examples in sponges, cnidarians, rotifers, nematodes, molluscs and arthropods which suggest that horizontal gene transfer in metazoans is not simply a curiosity. In addition, I stress the scarcity of studies in vertebrates and other animal groups and the importance of forthcoming studies to understand the importance and extent of horizontal gene transfer in animals. PMID:24403327

  20. Gene transfer for neovascular age-related macular degeneration.

    PubMed

    Campochiaro, Peter A

    2011-05-01

    Age-related macular degeneration (AMD) is a complex disease that has two phases: a degenerative phase often referred to as nonneovascular AMD (non-NVAMD) or dry AMD and a phase dominated by growth of new blood vessels in the subretinal space, referred to as NVAMD or wet AMD. Advances in the understanding of the molecular pathogenesis of NVAMD have led to new drug therapies that have provided major benefits to patients. However, those treatments require frequent intraocular injections that in many patients must be continued indefinitely to maintain visual benefits. Gene transfer to augment expression of endogenous antiangiogenic proteins is an alternative approach that has the potential to provide long-term stability in patients with NVAMD. Studies in animal models that mimic aspects of NVAMD have identified several possible transgenes, and a clinical trial in patients with advanced NVAMD has suggested that the approach may be feasible. Many important questions remain, but the rationale and preliminary data are compelling. The results of two ongoing clinical trials may answer several of the questions and help direct future research.

  1. HEAT TRANSFER AND TRITIUM PRODUCING SYSTEM

    DOEpatents

    Johnson, E.F.

    1962-06-01

    This invention related to a circulating lithium-containing blanket system in a neution source hav'ing a magnetic field associated therewith. The blanket serves simultaneously and efficiently as a heat transfer mediunm and as a source of tritium. The blanket is composed of a lithium-6-enriched fused salt selected from the group consisting of lithium nitrite, lithium nitrate, a mixture of said salts, a mixture of each of said salts with lithium oxide, and a mixture of said salts with each other and with lithium oxide. The moderator, which is contained within the blanket in a separate conduit, can be water. A stellarator is one of the neutron sources which can be used in this invention. (AEC)

  2. Infrared radiative energy transfer in gaseous systems

    NASA Technical Reports Server (NTRS)

    Tiwari, Surendra N.

    1991-01-01

    Analyses and numerical procedures are presented to investigate the radiative interactions in various energy transfer processes in gaseous systems. Both gray and non-gray radiative formulations for absorption and emission by molecular gases are presented. The gray gas formulations are based on the Planck mean absorption coefficient and the non-gray formulations are based on the wide band model correlations for molecular absorption. Various relations for the radiative flux and divergence of radiative flux are developed. These are useful for different flow conditions and physical problems. Specific plans for obtaining extensive results for different cases are presented. The procedure developed was applied to several realistic problems. Results of selected studies are presented.

  3. Gene recruitment--a common mechanism in the evolution of transfer RNA gene families.

    PubMed

    Wang, Xiujuan; Lavrov, Dennis V

    2011-04-01

    The evolution of alloacceptor transfer RNAs (tRNAs) has been traditionally thought to occur vertically and reflect the evolution of the genetic code. Yet there have been several indications that a tRNA gene could evolve horizontally, from a copy of an alloacceptor tRNA gene in the same genome. Earlier, we provided the first unambiguous evidence for the occurrence of such "tRNA gene recruitment" in nature--in the mitochondrial (mt) genome of the demosponge Axinella corrugata. Yet the extent and the pattern of this process in the evolution of tRNA gene families remained unclear. Here we analyzed tRNA genes from 21 mt genomes of demosponges as well as nuclear genomes of rhesus macaque, chimpanzee and human. We found four new cases of alloacceptor tRNA gene recruitment in mt genomes and eleven cases in the nuclear genomes. In most of these cases we observed a single nucleotide substitution at the middle position of the anticodon, which resulted in the change of not only the tRNA's amino-acid identity but also the class of the amino-acyl tRNA synthetases (aaRSs) involved in amino-acylation. We hypothesize that the switch to a different class of aaRSs may have prevented the conflict between anticodon and amino-acid identities of recruited tRNAs. Overall our results suggest that gene recruitment is a common phenomenon in tRNA multigene family evolution and should be taken into consideration when tRNA evolutionary history is reconstructed.

  4. Genomic Data Quality Impacts Automated Detection of Lateral Gene Transfer in Fungi

    PubMed Central

    Dupont, Pierre-Yves; Cox, Murray P.

    2017-01-01

    Lateral gene transfer (LGT, also known as horizontal gene transfer), an atypical mechanism of transferring genes between species, has almost become the default explanation for genes that display an unexpected composition or phylogeny. Numerous methods of detecting LGT events all rely on two fundamental strategies: primary structure composition or gene tree/species tree comparisons. Discouragingly, the results of these different approaches rarely coincide. With the wealth of genome data now available, detection of laterally transferred genes is increasingly being attempted in large uncurated eukaryotic datasets. However, detection methods depend greatly on the quality of the underlying genomic data, which are typically complex for eukaryotes. Furthermore, given the automated nature of genomic data collection, it is typically impractical to manually verify all protein or gene models, orthology predictions, and multiple sequence alignments, requiring researchers to accept a substantial margin of error in their datasets. Using a test case comprising plant-associated genomes across the fungal kingdom, this study reveals that composition- and phylogeny-based methods have little statistical power to detect laterally transferred genes. In particular, phylogenetic methods reveal extreme levels of topological variation in fungal gene trees, the vast majority of which show departures from the canonical species tree. Therefore, it is inherently challenging to detect LGT events in typical eukaryotic genomes. This finding is in striking contrast to the large number of claims for laterally transferred genes in eukaryotic species that routinely appear in the literature, and questions how many of these proposed examples are statistically well supported. PMID:28235827

  5. Identification of multiple independent horizontal gene transfers into poxviruses using a comparative genomics approach

    PubMed Central

    2008-01-01

    Background Poxviruses are important pathogens of humans, livestock and wild animals. These large dsDNA viruses have a set of core orthologs whose gene order is extremely well conserved throughout poxvirus genera. They also contain many genes with sequence and functional similarity to host genes which were probably acquired by horizontal gene transfer. Although phylogenetic trees can indicate the occurrence of horizontal gene transfer and even uncover multiple events, their use may be hampered by uncertainties in both the topology and the rooting of the tree. We propose to use synteny conservation around the horizontally transferred gene (HTgene) to distinguish between single and multiple events. Results Here we devise a method that incorporates comparative genomic information into the investigation of horizontal gene transfer, and we apply this method to poxvirus genomes. We examined the synteny conservation around twenty four pox genes that we identified, or which were reported in the literature, as candidate HTgenes. We found support for multiple independent transfers into poxviruses for five HTgenes. Three of these genes are known to be important for the survival of the virus in or out of the host cell and one of them increases susceptibility to some antiviral drugs. Conclusion In related genomes conserved synteny information can provide convincing evidence for multiple independent horizontal gene transfer events even in the absence of a robust phylogenetic tree for the HTgene. PMID:18304319

  6. A gene mapping expert system.

    PubMed

    Galland, J; Skolnick, M H

    1990-08-01

    Expert systems are now commonly developed to solve practical problems. Nevertheless, genetics has just begun to benefit from this new technology, since genetic expert systems are extremely rare and often purely experimental. A prototype for risk calculation in pedigrees was developed at the University of Utah, using a commercial frames/rules developmental shell (Intelligence Compiler), which runs on an IBM PC. When small data sets were used, the implementation functioned well, but it could not handle larger data sets. Performance became a major issue, with two possible solutions. The first possibility would have been to port the system to a more powerful machine, and the second would have been to use several different shells or languages, each efficiently representing a specific type of knowledge. Neither of these solutions was applicable in this case. From this experience, we learned that performance, portability, and modifiability were three major requirements for genetic expert systems. To achieve these goals, we implemented the gene mapping expert system GMES: (GMES is unrelated to the gene mapping system, GMS in Lisp combined with a frame/object shell (FROBS). We were able to efficiently represent, control, and optimize a gene mapping experiment, achieving portability by building GMES on top of a C-based version of Common Lisp. Lisp combined with the FROBS expert system shell permitted a declarative representation of each of the components of the experiment, resulting in a transplant specification of the problem within a maintainable system.

  7. Gene transfer to hemophilia A mice via oral delivery of FVIII-chitosan nanoparticles.

    PubMed

    Bowman, Katherine; Sarkar, Rita; Raut, Sanj; Leong, Kam W

    2008-12-18

    Effective oral delivery of a non-viral gene carrier would represent a novel and attractive strategy for therapeutic gene transfer. To evaluate the potential of this approach, we studied the oral gene delivery efficacy of DNA polyplexes composed of chitosan and Factor VIII DNA. Transgene DNA was detected in both local and systemic tissues following oral administration of the chitosan nanoparticles to hemophilia A mice. Functional factor VIII protein was detected in plasma by chromogenic and thrombin generation assays, reaching a peak level of 2-4% FVIII at day 22 after delivery. In addition, a bleeding challenge one month after DNA administration resulted in phenotypic correction in 13/20 mice given 250-600 microg of FVIII DNA in chitosan nanoparticles, compared to 1/13 mice given naked FVIII DNA and 0/6 untreated mice. While further optimization would be required to render this type of delivery system practical for hemophilia A gene therapy, the findings suggest the feasibility of oral, non-viral delivery for gene medicine applications.

  8. [Post-translational ligation and function of dual-vector transferred split CFTR gene].

    PubMed

    Zhu, Fu-Xiang; Liu, Ze-Long; Qu, Hui-Ge; Chi, Xiao-Yan

    2010-01-01

    The mutation of cystic fibrosis transmembrane conductance regulator (CFTR) gene leads to an autosomal recessive genetic disorder cystic fibrosis (CF). The gene therapy for CF using adeno-associated virus (AAV) vectors delivering CFTR gene is restricted by the contents limitation of AAV vectors. In this study the split CFTR genes severed at its regulatory domain were delivered by a dual-vector system with an intein-mediated protein trans-splicing as a technique to investigate the post-translational ligation of CFTR half proteins and its function as a chloride ion channel. A pair of eukaryotic expression vectors was constructed by breaking the human CFTR cDNA before Ser712 codon and fusing with Ssp DnaB intein coding sequences. After co-transfection into baby hamster kidney (BHK) cells followed by transient expression, patch clamps were carried out to record the chloride current of whole-cell and the activity of a single channel, and the ligation of two halves of CFTR was observed by Western blotting. The results showed that the intein-fused half genes co-transfected cells displayed a high whole cell chloride current and activity of a single channel indicating the functional recovery of chloride channel, and an intact CFTR protein band was figured out by CFTR-specific antibodies indicating that intein can efficiently ligate the separately expressed half CFTR proteins. The data demonstrated that protein splicing strategy could be used as a strategy in delivering CFTR gene by two vectors, encouraging our ongoing research program on dual AAV vector system based gene transfer in gene therapy for cystic fibrosis.

  9. Bagless Transfer System Welder Data Acquisition Software

    SciTech Connect

    Collins, Susan L.

    2003-09-30

    The Bagless Transfer System Welder Data Acquisition Software (BTS DAS) was developed by SRTC to replace a strip chart recorder that has been in place since the design of the BTS. During the welding process, critical weld parameters such as weld current and voltage, can give valuable information about the weld. In the past, weld data from the TID welding process, has been monitored using strip chart recorders. The data from the weld process, recorded on the strip chart recorder traces, are reviewed to analyze the weld. The BTS DAS improves this technology by digitizing the weld data which allows for automation of the analysis process. Also, the data files are now stored digitally as well as a hard copy printout, so they can be reanalyzed if needed. The BTS DAS performs the necessary functions to perform the data acquisition functions during the BTS Welding Process. It is important to monitor the critical weld parameters, current and voltage, during a weld as they can be used to set acceptance criteria for weld acceptance. The software monitors and records the weld current, voltage, and RPM data. The welder DAS is a passive device and does not control the welder. The BTS control system interfaces directly with the welder and the BTS DAS. Digital handshaking is used between the BTS DAS and the BTS control system to ensure that the DAS is ready to weld prior to allowing the operator to initiate the welding process.

  10. Gene Transfer Potential of Outer Membrane Vesicles of Acinetobacter baylyi and Effects of Stress on Vesiculation

    PubMed Central

    Fulsundar, Shweta; Harms, Klaus; Flaten, Gøril E.; Johnsen, Pål J.; Chopade, Balu Ananda

    2014-01-01

    Outer membrane vesicles (OMVs) are continually released from a range of bacterial species. Numerous functions of OMVs, including the facilitation of horizontal gene transfer (HGT) processes, have been proposed. In this study, we investigated whether OMVs contribute to the transfer of plasmids between bacterial cells and species using Gram-negative Acinetobacter baylyi as a model system. OMVs were extracted from bacterial cultures and tested for the ability to vector gene transfer into populations of Escherichia coli and A. baylyi, including naturally transformation-deficient mutants of A. baylyi. Anti-double-stranded DNA (anti-dsDNA) antibodies were used to determine the movement of DNA into OMVs. We also determined how stress affected the level of vesiculation and the amount of DNA in vesicles. OMVs were further characterized by measuring particle size distribution (PSD) and zeta potential. Transmission electron microscopy (TEM) and immunogold labeling were performed using anti-fluorescein isothiocyanate (anti-FITC)-conjugated antibodies and anti-dsDNA antibodies to track the movement of FITC-labeled and DNA-containing OMVs. Exposure to OMVs isolated from plasmid-containing donor cells resulted in HGT to A. baylyi and E. coli at transfer frequencies ranging from 10−6 to 10−8, with transfer efficiencies of approximately 103 and 102 per μg of vesicular DNA, respectively. Antibiotic stress was shown to affect the DNA content of OMVs as well as their hydrodynamic diameter and zeta potential. Morphological observations suggest that OMVs from A. baylyi interact with recipient cells in different ways, depending on the recipient species. Interestingly, the PSD measurements suggest that distinct size ranges of OMVs are released from A. baylyi. PMID:24657872

  11. Transfer functions of the conjugative integrating element pSAM2 from Streptomyces ambofaciens: characterization of a kil-kor system associated with transfer.

    PubMed

    Hagège, J; Pernodet, J L; Sezonov, G; Gerbaud, C; Friedmann, A; Guérineau, M

    1993-09-01

    pSAM2 is an 11-kb integrating element from Streptomyces ambofaciens. During matings, pSAM2 can be transferred at high frequency, forming pocks, which are zones of growth inhibition of the recipient strain. The nucleotide sequences of the regions involved in pSAM2 transfer, pock formation, and maintenance have been determined. Seven putative open reading frames with the codon usage typical of Streptomyces genes have been identified: traSA (306 amino acids [aa]), orf84 (84 aa), spdA (224 aa), spdB (58 aa), spdC (51 aa), spdD (104 aa), and korSA (259 aa). traSA is essential for pSAM2 intermycelial transfer and pock formation. It could encode a protein with similarities to the major transfer protein, Tra, of pIJ101. TraSA protein contains a possible nucleotide-binding sequence and a transmembrane segment. spdA, spdB, spdC, and spdD influence pock size and transfer efficiency and may be required for intramycelial transfer. A kil-kor system similar to that of pIJ101 is associated with pSAM2 transfer: the korSA (kil-override) gene product could control the expression of the traSA gene, which has lethal effects when unregulated (Kil phenotype). The KorSA protein resembles KorA of pIJ101 and repressor proteins belonging to the GntR family. Thus, the integrating element pSAM2 possesses for transfer general features of nonintegrating Streptomyces plasmids: different genes are involved in the different steps of the intermycelial and intramycelial transfer, and a kil-kor system is associated with transfer. However, some differences in the functional properties, organization, and sizes of the transfer genes compared with those of other Streptomyces plasmids have been found.

  12. Imprinted Genes and Satellite Loci Are Differentially Methylated in Bovine Somatic Cell Nuclear Transfer Clones

    PubMed Central

    Shen, Chih-Jie; Lin, Chiao-Chieh; Shen, Perng-Chih; Cheng, Winston T.K.; Chen, Hsiao-Ling; Chang, Tsung-Chou; Liu, Shyh-Shyan

    2013-01-01

    Abstract In mammals, genome-wide epigenetic reprogramming systems exist in primordial germ cells and zygotes. These reprogramming systems play crucial roles in regulating genome functions during critical stages of embryonic development, and they confer the stability of gene expression during mammalian development. The frequent unexpected loss of progeny from somatic cell nuclear transfer (SCNT) is an ongoing problem. In this study, we used six cloned bovines (named NT-1 to NT-6), which were created by ear fibroblast nuclear transfer and displayed short life spans with multiple organ defects, as an experimental model. We focus here on three imprinted genes (IGF2, H19, and XIST) and four satellite loci (Satellite I, Satellite II, Art2, and VNTR) to investigate their methylation changes. The results revealed that aberrant methylation frequently occurred in the analyzed imprinted genes, but not in the satellite loci, of the cloned bovines. After the bovine fibroblast cells were treated with the 5-aza-2(′)-deoxycytidine (5-Aza-dc) demethylation agent, the methylation percentages of the XIST and H19 putative differentially methylated region (DMR) were significantly decreased (XIST, p<0.01; H19, p<0.05) followed by an increase in their mRNA expression levels (p<0.01). Furthermore, we found that five short-lived cloned bovines (NT-1 to NT-5) exhibited more severe aberrant methylation changes in the three imprinted genes examined than the little longer-lived clone (NT-6) compared with wild-type (WT) cows. Our data suggest that the reprogramming of the methylation-controlled regions between the imprinted genes and satellite loci are differences and may be involved with additional mechanisms that need further elucidation. PMID:23961768

  13. Canister Transfer System Event Sequence Calculation

    SciTech Connect

    Richard Morissette

    2001-08-16

    The ''Department of Energy Spent Nuclear Fuel Canister, Transportation, and Monitored Geologic Repository Systems, Structures, and Components Performance Allocation Study'' (CRWMS M&O 2000b) allocated performance to both the canisters received at the Monitored Geologic Repository (MGR) and the MGR Canister Transfer System (CTS). The purpose of this calculation is to evaluate an assumed range of canister and CTS performance allocation failure probabilities and determine the effect of these failure probabilities on the frequency of a radionuclide release. Five canister types are addressed in this calculation; high-level radioactive waste (HLW) canisters containing vitrified borosilicate glass, HLW canisters containing immobilized plutonium surrounded by borosilicate glass (Pu/HLW canisters), Department of Energy (DOE) spent nuclear fuel (DSNF) standard canisters (4 sizes), DSNF multi-canister overpacks (MCOs) for N-reactor fuel and other selected DSNF, and naval spent nuclear fuel (SNF) canisters (2 sizes). The quality assurance program applies to this calculation, and the work is performed in accordance with procedure AP-3.12Q, ''Calculations''. The work done for this calculation was evaluated according to AP-2.21Q, ''Quality Determinations and Planning for Scientific, Engineering, and Regulatory Compliance Activities'' that determined this activity to be subject to the requirements of DOE/RW-0333P, ''Quality Assurance Requirements and Description'' (DOE 2000a). This work was performed in accordance with the ''Technical Work Plan for: Department of Energy Nuclear Fuel Work Packages'' (CRWMS M&O 2000c) for this activity.

  14. Space Biosensor Systems: Implications for Technology Transfer

    NASA Technical Reports Server (NTRS)

    Hines, J. W.; Somps, C. J.; Madou, M.; Imprescia, Clifford C. (Technical Monitor)

    1997-01-01

    To meet the need for continuous, automated monitoring of animal subjects, including; humans, during space flight, NASA is developing advanced physiologic sensor and biotelemetry system technologies. The ability to continuously track basic physiological parameters, such as heart rate, blood pH, and body temperature, in untethered subjects in space is a challenging task. At NASA's Ames Research Center, where a key focus is gravitational biology research, engineers have teamed with life scientists to develop wireless sensor systems for automated physiologic monitoring of animal models as small as the rat. This technology is also being adapted, in collaboration with medical professionals, to meet human clinical monitoring needs both in space and on the ground. Thus, these advanced monitoring technologies have important dual-use functions; they meet space flight data collection requirements and constraints, while concurrently addressing a number of monitoring and data acquisition challenges on the ground in areas of clinical monitoring and biomedical research. Additional applications for these and related technologies are being sought and additional partnerships established that enhance development efforts, reduce costs and facilitate technology infusion between the public and private sectors. This paper describes technology transfer and co-development projects that have evolved out of NASA's miniaturized, implantable chemical sensor development efforts.

  15. Proteomic profiling of salivary gland after nonviral gene transfer mediated by conventional plasmids and minicircles

    PubMed Central

    Geguchadze, Ramaz; Wang, Zhimin; Zourelias, Lee; Perez-Riveros, Paola; Edwards, Paul C; Machen, Laurie; Passineau, Michael J

    2014-01-01

    In this study, we compared gene transfer efficiency and host response to ultrasound-assisted, nonviral gene transfer with a conventional plasmid and a minicircle vector in the submandibular salivary glands of mice. Initially, we looked at gene transfer efficiency with equimolar amounts of the plasmid and minicircle vectors, corroborating an earlier report showing that minicircle is more efficient in the context of a physical method of gene transfer. We then sought to characterize the physiological response of the salivary gland to exogenous gene transfer using global proteomic profiling. Somewhat surprisingly, we found that sonoporation alone, without a gene transfer vector present, had virtually no effect on the salivary gland proteome. However, when a plasmid vector was used, we observed profound perturbations of the salivary gland proteome that compared in magnitude to that seen in a previous report after high doses of adeno-associated virus. Finally, we found that gene transfer with a minicircle induces only minor proteomic alterations that were similar to sonoporation alone. Using mass spectrometry, we assigned protein IDs to 218 gel spots that differed between plasmid and minicircle. Bioinformatic analysis of these proteins demonstrated convergence on 68 known protein interaction pathways, most notably those associated with innate immunity, cellular stress, and morphogenesis. PMID:25414909

  16. Apramycin resistance as a selective marker for gene transfer in mycobacteria.

    PubMed Central

    Paget, E; Davies, J

    1996-01-01

    We have explored the potential of using the apramycin resistance gene as a marker in mycobacterial gene transfer studies. Shuttle plasmids available for both electroporation and conjugation studies have been constructed, and we have successfully validated the use of the apramycin resistance gene as a component of cloning vectors for Mycobacterium smegmatis, M. bovis BCG, and M. tuberculosis. PMID:8892841

  17. Major role of local immune responses in antibody formation to factor IX in AAV gene transfer.

    PubMed

    Wang, L; Cao, O; Swalm, B; Dobrzynski, E; Mingozzi, F; Herzog, R W

    2005-10-01

    The risk of an immune response to the coagulation factor IX (F.IX) transgene product is a concern in gene therapy for the X-linked bleeding disorder hemophilia B. In order to investigate the mechanism of F.IX-specific lymphocyte activation in the context of adeno-associated viral (AAV) gene transfer to skeletal muscle, we injected AAV-2 vector expressing human F.IX (hF.IX) into outbred immune-competent mice. Systemic hF.IX levels were transiently detected in the circulation, but diminished concomitant with activation of CD4+ T and B cells. ELISPOT assays documented robust responses to hF.IX in the draining lymph nodes of injected muscle by day 14. Formation of inhibitory antibodies to hF.IX was observed over a wide range of vector doses, with increased doses causing stronger immune responses. A prolonged inflammatory reaction in muscle started at 1.5-2 months, but ultimately failed to eliminate transgene expression. By 1.5 months, hF.IX antigen re-emerged in circulation in approximately 70% of animals injected with high vector dose. Hepatic gene transfer elicited only infrequent and weaker immune responses, with higher vector doses causing a reduction in T-cell responses to hF.IX. In summary, the data document substantial influence of target tissue, local antigen presentation, and antigen levels on lymphocyte responses to F.IX.

  18. Horizontal Gene Transfer of Pectinases from Bacteria Preceded the Diversification of Stick and Leaf Insects

    PubMed Central

    Shelomi, Matan; Danchin, Etienne G. J.; Heckel, David; Wipfler, Benjamin; Bradler, Sven; Zhou, Xin; Pauchet, Yannick

    2016-01-01

    Genes acquired by horizontal transfer are increasingly being found in animal genomes. Understanding their origin and evolution requires knowledge about the phylogenetic relationships from both source and recipient organisms. We used RNASeq data and respective assembled transcript libraries to trace the evolutionary history of polygalacturonase (pectinase) genes in stick insects (Phasmatodea). By mapping the distribution of pectinase genes on a Polyneoptera phylogeny, we identified the transfer of pectinase genes from known phasmatodean gut microbes into the genome of an early euphasmatodean ancestor that took place between 60 and 100 million years ago. This transfer preceded the rapid diversification of the suborder, enabling symbiont-free pectinase production that would increase the insects’ digestive efficiency and reduce dependence on microbes. Bacteria-to-insect gene transfer was thought to be uncommon, however the increasing availability of large-scale genomic data may change this prevailing notion. PMID:27210832

  19. Gene Transfer Efficiency in Gonococcal Biofilms: Role of Biofilm Age, Architecture, and Pilin Antigenic Variation

    PubMed Central

    Kouzel, Nadzeya; Oldewurtel, Enno R.

    2015-01-01

    ABSTRACT Extracellular DNA is an important structural component of many bacterial biofilms. It is unknown, however, to which extent external DNA is used to transfer genes by means of transformation. Here, we quantified the acquisition of multidrug resistance and visualized its spread under selective and nonselective conditions in biofilms formed by Neisseria gonorrhoeae. The density and architecture of the biofilms were controlled by microstructuring the substratum for bacterial adhesion. Horizontal transfer of antibiotic resistance genes between cocultured strains, each carrying a single resistance, occurred efficiently in early biofilms. The efficiency of gene transfer was higher in early biofilms than between planktonic cells. It was strongly reduced after 24 h and independent of biofilm density. Pilin antigenic variation caused a high fraction of nonpiliated bacteria but was not responsible for the reduced gene transfer at later stages. When selective pressure was applied to dense biofilms using antibiotics at their MIC, the double-resistant bacteria did not show a significant growth advantage. In loosely connected biofilms, the spreading of double-resistant clones was prominent. We conclude that multidrug resistance readily develops in early gonococcal biofilms through horizontal gene transfer. However, selection and spreading of the multiresistant clones are heavily suppressed in dense biofilms. IMPORTANCE Biofilms are considered ideal reaction chambers for horizontal gene transfer and development of multidrug resistances. The rate at which genes are exchanged within biofilms is unknown. Here, we quantified the acquisition of double-drug resistance by gene transfer between gonococci with single resistances. At early biofilm stages, the transfer efficiency was higher than for planktonic cells but then decreased with biofilm age. The surface topography affected the architecture of the biofilm. While the efficiency of gene transfer was independent of the

  20. British Columbia/Alberta Transfer System Protocol

    ERIC Educational Resources Information Center

    British Columbia Council on Admissions and Transfer, 2007

    2007-01-01

    The purpose of this agreement is to provide assurance to students that they will receive transfer credit for courses or programs they have successfully completed where the content/outcomes are demonstrably equivalent to those offered at the institution to which they transfer. This protocol is undertaken by the British Columbia Council on…

  1. 40 CFR 63.689 - Standards: Transfer systems.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ....689 Standards: Transfer systems. (a) The provisions of this section apply to the control of air... section for such air emission control. (b) For each transfer system that is subject to this section and is an individual drain system, the owner or operator shall control air emissions in accordance with...

  2. 40 CFR 63.689 - Standards: Transfer systems.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ....689 Standards: Transfer systems. (a) The provisions of this section apply to the control of air... section for such air emission control. (b) For each transfer system that is subject to this section and is an individual drain system, the owner or operator shall control air emissions in accordance with...

  3. Horizontal gene transfer and the evolution of transcriptionalregulation in Escherichia coli

    SciTech Connect

    Price, Morgan N.; Dehal, Paramvir S.; Arkin, Adam P.

    2007-12-20

    Background: Most bacterial genes were acquired by horizontalgene transfer from other bacteria instead of being inherited bycontinuous vertical descent from an ancient ancestor}. To understand howthe regulation of these {acquired} genes evolved, we examined theevolutionary histories of transcription factors and of regulatoryinteractions from the model bacterium Escherichia coli K12. Results:Although most transcription factors have paralogs, these usually arose byhorizontal gene transfer rather than by duplication within the E. colilineage, as previously believed. In general, most neighbor regulators --regulators that are adjacent to genes that they regulate -- were acquiredby horizontal gene transfer, while most global regulators evolvedvertically within the gamma-Proteobacteria. Neighbor regulators wereoften acquired together with the adjacent operon that they regulate, sothe proximity might be maintained by repeated transfers (like "selfishoperons"). Many of the as-yet-uncharacterized (putative) regulators havealso been acquired together with adjacent genes, so we predict that theseare neighbor regulators as well. When we analyzed the histories ofregulatory interactions, we found that the evolution of regulation byduplication was rare, and surprisingly, many of the regulatoryinteractions that are shared between paralogs result from convergentevolution. Another surprise was that horizontally transferred genes aremore likely than other genes to be regulated by multiple regulators, andmost of this complex regulation probably evolved after the transfer.Conclusions: Our results highlight the rapid evolution of niche-specificgene regulation in bacteria.

  4. Use of HIV as a gene transfer vector.

    PubMed

    Pluta, Krzysztof; Kacprzak, Magdalena Marta

    2009-01-01

    Despite the extensive research efforts over the past 25 years that have focused on HIV, there is still no cure for AIDS. However, tremendous progress in the understanding of the structure and biology of the HIV virus led to the development of safe and potent HIV-based transgene delivery vectors. These genetic vehicles are referred to as lentiviral vectors. They appear to be better suited for particular applications, such as transgene delivery into stem cells, compared to other viral- and non-viral vectors. This is because Lentivirus-based vectors can efficiently infect nondividing and slowly dividing cells. In the present review article, the current state of understanding of HIV-1 is discussed and the main characteristics that had an impact on vector design are outlined. A historical view on the vector concept is presented to facilitate discussion of recent results in vector engineering in a broader context. Subsequently, a state of the art overview concerning vector construction and vector production is given. This review also touches upon the subject of lentiviral vector safety and related topics that can be helpful in addressing this issue are discussed. Finally, examples of Lentivirus-based gene delivery systems and their applications are presented, with emphasis on animal transgenesis and human gene therapy.

  5. Safety and efficacy of gene transfer for Leber's congenital amaurosis.

    PubMed

    Maguire, Albert M; Simonelli, Francesca; Pierce, Eric A; Pugh, Edward N; Mingozzi, Federico; Bennicelli, Jeannette; Banfi, Sandro; Marshall, Kathleen A; Testa, Francesco; Surace, Enrico M; Rossi, Settimio; Lyubarsky, Arkady; Arruda, Valder R; Konkle, Barbara; Stone, Edwin; Sun, Junwei; Jacobs, Jonathan; Dell'Osso, Lou; Hertle, Richard; Ma, Jian-xing; Redmond, T Michael; Zhu, Xiaosong; Hauck, Bernd; Zelenaia, Olga; Shindler, Kenneth S; Maguire, Maureen G; Wright, J Fraser; Volpe, Nicholas J; McDonnell, Jennifer Wellman; Auricchio, Alberto; High, Katherine A; Bennett, Jean

    2008-05-22

    Leber's congenital amaurosis (LCA) is a group of inherited blinding diseases with onset during childhood. One form of the disease, LCA2, is caused by mutations in the retinal pigment epithelium-specific 65-kDa protein gene (RPE65). We investigated the safety of subretinal delivery of a recombinant adeno-associated virus (AAV) carrying RPE65 complementary DNA (cDNA) (ClinicalTrials.gov number, NCT00516477 [ClinicalTrials.gov]). Three patients with LCA2 had an acceptable local and systemic adverse-event profile after delivery of AAV2.hRPE65v2. Each patient had a modest improvement in measures of retinal function on subjective tests of visual acuity. In one patient, an asymptomatic macular hole developed, and although the occurrence was considered to be an adverse event, the patient had some return of retinal function. Although the follow-up was very short and normal vision was not achieved, this study provides the basis for further gene therapy studies in patients with LCA.

  6. Plasmid transfer by conjugation as a possible route of horizontal gene transfer and recombination in Xylella fastidiosa

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Horizontal gene transfer is an important component of evolution and adaptation of bacterial species. Xylella fastidiosa has the ability to incorporate exogenous DNA into its genome by homologous recombination at relatively high rates. This genetic recombination is believed to play a role in adaptati...

  7. Horizontally transferred genes in the genome of Pacific white shrimp, Litopenaeus vannamei

    PubMed Central

    2013-01-01

    Background In recent years, as the development of next-generation sequencing technology, a growing number of genes have been reported as being horizontally transferred from prokaryotes to eukaryotes, most of them involving arthropods. As a member of the phylum Arthropoda, the Pacific white shrimp Litopenaeus vannamei has to adapt to the complex water environments with various symbiotic or parasitic microorganisms, which provide a platform for horizontal gene transfer (HGT). Results In this study, we analyzed the genome-wide HGT events in L. vannamei. Through homology search and phylogenetic analysis, followed by experimental PCR confirmation, 14 genes with HGT event were identified: 12 of them were transferred from bacteria and two from fungi. Structure analysis of these genes showed that the introns of the two fungi-originated genes were substituted by shrimp DNA fragment, two genes transferred from bacteria had shrimp specific introns inserted in them. Furthermore, around other three bacteria-originated genes, there were three large DNA segments inserted into the shrimp genome. One segment was a transposon that fully transferred, and the other two segments contained only coding regions of bacteria. Functional prediction of these 14 genes showed that 6 of them might be related to energy metabolism, and 4 others related to defense of the organism. Conclusions HGT events from bacteria or fungi were happened in the genome of L. vannamei, and these horizontally transferred genes can be transcribed in shrimp. This is the first time to report the existence of horizontally transferred genes in shrimp. Importantly, most of these genes are exposed to a negative selection pressure and appeared to be functional. PMID:23914989

  8. A new computational method for the detection of horizontal gene transfer events.

    PubMed

    Tsirigos, Aristotelis; Rigoutsos, Isidore

    2005-01-01

    In recent years, the increase in the amounts of available genomic data has made it easier to appreciate the extent by which organisms increase their genetic diversity through horizontally transferred genetic material. Such transfers have the potential to give rise to extremely dynamic genomes where a significant proportion of their coding DNA has been contributed by external sources. Because of the impact of these horizontal transfers on the ecological and pathogenic character of the recipient organisms, methods are continuously sought that are able to computationally determine which of the genes of a given genome are products of transfer events. In this paper, we introduce and discuss a novel computational method for identifying horizontal transfers that relies on a gene's nucleotide composition and obviates the need for knowledge of codon boundaries. In addition to being applicable to individual genes, the method can be easily extended to the case of clusters of horizontally transferred genes. With the help of an extensive and carefully designed set of experiments on 123 archaeal and bacterial genomes, we demonstrate that the new method exhibits significant improvement in sensitivity when compared to previously published approaches. In fact, it achieves an average relative improvement across genomes of between 11 and 41% compared to the Codon Adaptation Index method in distinguishing native from foreign genes. Our method's horizontal gene transfer predictions for 123 microbial genomes are available online at http://cbcsrv.watson.ibm.com/HGT/.

  9. Cellular Immune Response Against Firefly Luciferase After Sleeping Beauty–Mediated Gene Transfer In Vivo

    PubMed Central

    Podetz-Pedersen, Kelly M.; Vezys, Vaiva; Somia, Nikunj V.; Russell, Stephen J.

    2014-01-01

    Abstract The Sleeping Beauty (SB) transposon system has been shown to mediate new gene sequence integration resulting in long-term expression. Here the effectiveness of hyperactive SB100X transposase was tested, and we found that hydrodynamic co-delivery of a firefly luciferase transposon (pT2/CaL) along with SB100X transposase (pCMV-SB100X) resulted in remarkably sustained, high levels of luciferase expression. However, after 4 weeks there was a rapid, animal-by-animal loss of luciferase expression that was not observed in immunodeficient mice. We hypothesized that this sustained, high-level luciferase expression achieved using the SB100X transposase elicits an immune response in pT2/CaL co-administered mice, which was supported by the rapid loss of luciferase expression upon challenge of previously treated animals and in naive animals adoptively transferred with splenocytes from previously treated animals. Specificity of the immune response to luciferase was demonstrated by increased cytokine expression in splenocytes after exposure to luciferase peptide in parallel with MHC I–luciferase peptide tetramer binding. This anti-luciferase immune response observed following continuous, high-level luciferase expression in vivo clearly impacts its use as an in vivo reporter. As both an immunogen and an extremely sensitive reporter, luciferase is also a useful model system for the study of immune responses following in vivo gene transfer and expression. PMID:25093708

  10. W-314, waste transfer alternative piping system description

    SciTech Connect

    Papp, I.G.

    1998-04-30

    It is proposed that the reliability, operability, and flexibility of the Retrieval Transfer System be substantially upgraded by replacing the planned single in-farm pipeline from the AN-AY-AZ-(SY) Tank Farm Complex to the AP Farm with three parallel pipelines outside the tank farms. The proposed system provides simplified and redundant routes for the various transfer missions, and prevents the risk of transfer gridlock when the privatization effort swings into full operation.

  11. Bacteriophage WO Can Mediate Horizontal Gene Transfer in Endosymbiotic Wolbachia Genomes

    PubMed Central

    Wang, Guan H.; Sun, Bao F.; Xiong, Tuan L.; Wang, Yan K.; Murfin, Kristen E.; Xiao, Jin H.; Huang, Da W.

    2016-01-01

    Phage-mediated horizontal gene transfer (HGT) is common in free-living bacteria, and many transferred genes can play a significant role in their new bacterial hosts. However, there are few reports concerning phage-mediated HGT in endosymbionts (obligate intracellular bacteria within animal or plant hosts), such as Wolbachia. The Wolbachia-infecting temperate phage WO can actively shift among Wolbachia genomes and has the potential to mediate HGT between Wolbachia strains. In the present study, we extend previous findings by validating that the phage WO can mediate transfer of non-phage genes. To do so, we utilized bioinformatic, phylogenetic, and molecular analyses based on all sequenced Wolbachia and phage WO genomes. Our results show that the phage WO can mediate HGT between Wolbachia strains, regardless of whether the transferred genes originate from Wolbachia or other unrelated bacteria. PMID:27965627

  12. SAFETY AND EFFICIENCY OF MODULATING PARACELLULAR PERMEABILITY TO ENHANCE AIRWAY EPITHELIAL GENE TRANSFER IN VIVO

    EPA Science Inventory


    ABSTRACT

    We evaluated the safety of agents that enhance gene transfer by modulating paracellular permeability. Lactate dehydrogenase (LDH) and cytokine release were measured in polarized primary human airway epithelial (HAE) cells after luminal application of vehicle, ...

  13. Genome-wide identification of horizontal gene transfer in Fusarium verticillioides

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Horizontal gene transfer (HGT), the exchange and stable integration of genetic material between different lineages, breaks species boundaries and generates new biological diversity. In eukaryotes, despite potential barriers, like the nuclear envelope and multicellularity, HGT may be facilitated by t...

  14. An Examination of Learning Transfer System Characteristics across Organizational Settings

    ERIC Educational Resources Information Center

    Holton, Elwood F., III; Chen, Hsin-Chih; Naquin, Sharon S.

    2003-01-01

    No previous research in the United States has compared and contrasted learning transfer systems across organizations and training types, primarily because no standard instruments were used in previous research. This study, based on a subset of responses in the Learning Transfer System Inventory response database, is the first to conduct such a…

  15. Genome-Wide Survey of Gut Fungi (Harpellales) Reveals the First Horizontally Transferred Ubiquitin Gene from a Mosquito Host

    PubMed Central

    Wang, Yan; White, Merlin M.; Kvist, Sebastian; Moncalvo, Jean-Marc

    2016-01-01

    Harpellales, an early-diverging fungal lineage, is associated with the digestive tracts of aquatic arthropod hosts. Concurrent with the production and annotation of the first four Harpellales genomes, we discovered that Zancudomyces culisetae, one of the most widely distributed Harpellales species, encodes an insect-like polyubiquitin chain. Ubiquitin and ubiquitin-like proteins are universally involved in protein degradation and regulation of immune response in eukaryotic organisms. Phylogenetic analyses inferred that this polyubiquitin variant has a mosquito origin. In addition, its amino acid composition, animal-like secondary structure, as well as the fungal nature of flanking genes all further support this as a horizontal gene transfer event. The single-copy polyubiquitin gene from Z. culisetae has lower GC ratio compared with homologs of insect taxa, which implies homogenization of the gene since its putatively ancient transfer. The acquired polyubiquitin gene may have served to improve important functions within Z. culisetae, by perhaps exploiting the insect hosts’ ubiquitin-proteasome systems in the gut environment. Preliminary comparisons among the four Harpellales genomes highlight the reduced genome size of Z. culisetae, which corroborates its distinguishable symbiotic lifestyle. This is the first record of a horizontally transferred ubiquitin gene from disease-bearing insects to the gut-dwelling fungal endobiont and should invite further exploration in an evolutionary context. PMID:27343289

  16. A statistical model for bacterial speciation triggered by lateral gene transfer

    NASA Astrophysics Data System (ADS)

    Sidhu, Sunjeet; Peng, Wequin

    2006-03-01

    The process of bacterial speciation has been a major unresolved issue in the study of bacterial evolution. It has been proposed that lateral gene transfer and homologous recombination play critical and complementary roles in speciation. We introduce a statistical model, of a population, for the evolution under lateral gene transfer and local homologous recombination. We examine the evolutionary dynamics and its dependence on various evolutionary operators. J. G. Lawrence, Theor. Popul. Biol. 61, 449(2002).

  17. Combining Genes from Multiple Phages for Improved Cell Lysis and DNA Transfer from Escherichia coli to Bacillus subtilis

    PubMed Central

    Juhas, Mario; Wong, Christine; Ajioka, James W.

    2016-01-01

    The ability to efficiently and reliably transfer genetic circuits between the key synthetic biology chassis, such as Escherichia coli and Bacillus subtilis, constitutes one of the major hurdles of the rational genome engineering. Using lambda Red recombineering we integrated the thermosensitive lambda repressor and the lysis genes of several bacteriophages into the E. coli chromosome. The lysis of the engineered autolytic cells is inducible by a simple temperature shift. We improved the lysis efficiency by introducing different combinations of lysis genes from bacteriophages lambda, ΦX174 and MS2 under the control of the thermosensitive lambda repressor into the E. coli chromosome. We tested the engineered autolytic cells by transferring plasmid and bacterial artificial chromosome (BAC)-borne genetic circuits from E. coli to B. subtilis. Our engineered system combines benefits of the two main synthetic biology chassis, E. coli and B. subtilis, and allows reliable and efficient transfer of DNA edited in E. coli into B. subtilis. PMID:27798678

  18. An Approach for Treating the Hepatobiliary Disease of Cystic Fibrosis by Somatic Gene Transfer

    NASA Astrophysics Data System (ADS)

    Yang, Yiping; Raper, Steven E.; Cohn, Jonathan A.; Engelhardt, John F.; Wilson, James M.

    1993-05-01

    Cystic fibrosis (CF) is an inherited disease of epithelial cell ion transport that is associated with pathology in multiple organ systems, including lung, pancreas, and liver. As treatment of the pulmonary manifestations of CF has improved, management of CF liver disease has become increasingly important in adult patients. This report describes an approach for treating CF liver disease by somatic gene transfer. In situ hybridization and immunocytochemistry analysis of rat liver sections indicated that the endogenous CFTR (cystic fibrosis transmembrane conductance regulator) gene is primarily expressed in the intrahepatic biliary epithelial cells. To specifically target recombinant genes to the biliary epithelium in vivo, recombinant adenoviruses expressing lacZ or human CFTR were infused retrograde into the biliary tract through the common bile duct. Conditions were established for achieving recombinant gene expression in virtually all cells of the intrahepatic bile ducts in vivo. Expression persisted in the smaller bile ducts for the duration of the experiment, which was 21 days. These studies suggest that it may be feasible to prevent CF liver disease by genetically reconstituting CFTR expression in the biliary tract, using an approach that is clinically feasible.

  19. Efficient Non-Viral Ocular Gene Transfer with Compacted DNA Nanoparticles

    PubMed Central

    Farjo, Rafal; Skaggs, Jeff; Quiambao, Alexander B.; Cooper, Mark J.; Naash, Muna I.

    2006-01-01

    Background The eye is an excellent candidate for gene therapy as it is immune privileged and much of the disease-causing genetics are well understood. Towards this goal, we evaluated the efficiency of compacted DNA nanoparticles as a system for non-viral gene transfer to ocular tissues. The compacted DNA nanoparticles examined here have been shown to be safe and effective in a human clinical trial, have no theoretical limitation on plasmid size, do not provoke immune responses, and can be highly concentrated. Methods and Findings Here we show that these nanoparticles can be targeted to different tissues within the eye by varying the site of injection. Almost all cell types of the eye were capable of transfection by the nanoparticle and produced robust levels of gene expression that were dose-dependent. Most impressively, subretinal delivery of these nanoparticles transfected nearly all of the photoreceptor population and produced expression levels almost equal to that of rod opsin, the highest expressed gene in the retina. Conclusions As no deleterious effects on retinal function were observed, this treatment strategy appears to be clinically viable and provides a highly efficient non-viral technology to safely deliver and express nucleic acids in the retina and other ocular tissues. PMID:17183666

  20. Immune deficiency enhances expression of recombinant human antibody in mice after nonviral in vivo gene transfer.

    PubMed

    Kitaguchi, Kohji; Toda, Mikako; Takekoshi, Masataka; Maeda, Fumiko; Muramatsu, Tatsuo; Murai, Atsushi

    2005-10-01

    A cDNA encoding human antibody against hepatitis B virus was expressed in normal and severe combined immune deficiency (SCID) mice to clarify whether or not host immune status affects circulating levels of the recombinant human antibody (RhAb) after nonviral in vivo gene transfer. For transferring genes, either electroporation (EP) or hydrodynamics-based transfection (HD) was employed. The former was applied to the leg muscle to express the gene, while the latter primarily targeted foreign gene expression in the liver. The expressed RhAb was secreted into the blood circulation, and its existence was assayed by ELISA. Prior to the investigation of host immune status, suitable forms of plasmid expression vectors and types of electrodes were determined in normal mice. Results showed that the vector encoding both the light and heavy chains driven by the CMV promoter had the highest plasma RhAb concentrations, and a pair of pincette-type electrodes conferred the best performance. In both EP and HD, the SCID state showed an increased and prolonged RhAb production in the blood circulation due probably to suppressed recognition of RhAb as a foreign protein to the host animal. The difference in gene transfer methods demonstrated a characteristic pattern: an early and sharp rise followed by a relatively rapid decrease in HD, in contrast to a gradual rise followed by a plateau level maintained in EP. As a result, with the same amount of gene transferred, the plasma RhAb concentrations for the first 7 or 8 weeks were higher in HD than EP, while the reverse was true for the latter period. Multiple gene transfer contributed to maintaining and prolonging high RhAb concentrations in plasma by both methods with similar characteristic patterns accompanying the respective gene transfer method. These results suggest the importance of host immunological potency for maintaining plasma RhAb concentrations if these gene transfer technologies are used for clinical and therapeutic purposes.

  1. Recombinant AAV-mediated gene transfer to the retina: gene therapy perspectives.

    PubMed

    Rolling, F

    2004-10-01

    Retinal degenerative diseases such as retinal macular degeneration and retinitis pigmentosa constitute a broad group of diseases that all share one critical feature, the progressive apoptotic loss of cells in the retina. There is currently no effective treatment available by which the course of these disorders can be modified, and visual dysfunction often progresses to total blindness. Gene therapy represents an attractive approach to treating retinal degeneration because the eye is easily accessible and allows local application of therapeutic vectors with reduced risk of systemic effects. Furthermore, transgene expression within the retina and effects of treatments may be monitored by a variety of noninvasive examinations. An increasing number of strategies for molecular treatment of retinal disease rely on recombinant adeno-associated virus (rAAV) as a therapeutic gene delivery vector. Before rAAV-mediated gene therapy for retinal degeneration becomes a reality, there are a number of important requirements that include: (1) evaluation of different rAAV serotypes, (2) screening of vectors in large animals in order to ensure that they mediate safe and long-term gene expression, (3) appropriate regulation of therapeutic gene expression, (4) evaluation of vectors carrying a therapeutic gene in relevant animal models, (5) identification of suitable patients, and finally (6) manufacture of clinical grade vector. All these steps towards gene therapy are still being explored. Outcomes of these studies will be discussed in the order in which they occur, from vector studies to preclinical assessment of the therapeutic potential of rAAV in animal models of retinal degeneration.

  2. An XMRV Derived Retroviral Vector as a Tool for Gene Transfer

    PubMed Central

    2011-01-01

    Background Retroviral vectors are widely used tools for gene delivery and gene therapy. They are useful for gene expression studies and genetic manipulation in vitro and in vivo. Many retroviral vectors are derived from the mouse gammaretrovirus, murine leukemia virus (MLV). These vectors have been widely used in gene therapy clinical trials. XMRV, initially found in prostate cancer tissue, was the first human gammaretrovirus described. Findings We developed a new retroviral vector based on XMRV called pXC. It was developed for gene transfer to human cells and is produced by transient cotransfection of LNCaP cells with pXC and XMRV-packaging plasmids. Conclusions We demonstrated that pXC mediates expression of inserted transgenes in cell lines. This new vector will be a useful tool for gene transfer in human and non-human cell lines, including gene therapy studies. PMID:21651801

  3. Direct Gene Transfer into Human Cultured Cells Facilitated by Laser Micropuncture of the Cell Membrane

    NASA Astrophysics Data System (ADS)

    Tao, Wen; Wilkinson, Joyce; Stanbridge, Eric J.; Berns, Michael W.

    1987-06-01

    The selective alteration of the cellular genome by laser microbeam irradiation has been extensively applied in cell biology. We report here the use of the third harmonic (355 nm) of an yttrium-aluminum garnet laser to facilitate the direct transfer of the neo gene into cultured human HT1080-6TG cells. The resultant transformants were selected in medium containing an aminoglycoside antibiotic, G418. Integration of the neo gene into individual human chromosomes and expression of the gene were demonstrated by Southern blot analyses, microcell-mediated chromosome transfer, and chromosome analyses. The stability of the integrated neo gene in the transformants was shown by a comparative growth assay in selective and nonselective media. Transformation and incorporation of the neo gene into the host genome occurred at a frequency of 8 × 10-4-3 × 10-3. This method appears to be 100-fold more efficient than the standard calcium phosphate-mediated method of DNA transfer.

  4. Horizontal gene transfer from diverse bacteria to an insect genome enables a tripartite nested mealybug symbiosis.

    PubMed

    Husnik, Filip; Nikoh, Naruo; Koga, Ryuichi; Ross, Laura; Duncan, Rebecca P; Fujie, Manabu; Tanaka, Makiko; Satoh, Nori; Bachtrog, Doris; Wilson, Alex C C; von Dohlen, Carol D; Fukatsu, Takema; McCutcheon, John P

    2013-06-20

    The smallest reported bacterial genome belongs to Tremblaya princeps, a symbiont of Planococcus citri mealybugs (PCIT). Tremblaya PCIT not only has a 139 kb genome, but possesses its own bacterial endosymbiont, Moranella endobia. Genome and transcriptome sequencing, including genome sequencing from a Tremblaya lineage lacking intracellular bacteria, reveals that the extreme genomic degeneracy of Tremblaya PCIT likely resulted from acquiring Moranella as an endosymbiont. In addition, at least 22 expressed horizontally transferred genes from multiple diverse bacteria to the mealybug genome likely complement missing symbiont genes. However, none of these horizontally transferred genes are from Tremblaya, showing that genome reduction in this symbiont has not been enabled by gene transfer to the host nucleus. Our results thus indicate that the functioning of this three-way symbiosis is dependent on genes from at least six lineages of organisms and reveal a path to intimate endosymbiosis distinct from that followed by organelles.

  5. Inter-genomic displacement via lateral gene transfer of bacterial trp operons in an overall context of vertical genealogy

    PubMed Central

    Xie, Gary; Bonner, Carol A; Song, Jian; Keyhani, Nemat O; Jensen, Roy A

    2004-01-01

    Background The growing conviction that lateral gene transfer plays a significant role in prokaryote genealogy opens up a need for comprehensive evaluations of gene-enzyme systems on a case-by-case basis. Genes of tryptophan biosynthesis are frequently organized as whole-pathway operons, an attribute that is expected to facilitate multi-gene transfer in a single step. We have asked whether events of lateral gene transfer are sufficient to have obscured our ability to track the vertical genealogy that underpins tryptophan biosynthesis. Results In 47 complete-genome Bacteria, the genes encoding the seven catalytic domains that participate in primary tryptophan biosynthesis were distinguished from any paralogs or xenologs engaged in other specialized functions. A reliable list of orthologs with carefully ascertained functional roles has thus been assembled and should be valuable as an annotation resource. The protein domains associated with primary tryptophan biosynthesis were then concatenated, yielding single amino-acid sequence strings that represent the entire tryptophan pathway. Lateral gene transfer of several whole-pathway trp operons was demonstrated by use of phylogenetic analysis. Lateral gene transfer of partial-pathway trp operons was also shown, with newly recruited genes functioning either in primary biosynthesis (rarely) or specialized metabolism (more frequently). Conclusions (i) Concatenated tryptophan protein trees are congruent with 16S rRNA subtrees provided that the genomes represented are of sufficiently close phylogenetic spacing. There are currently seven tryptophan congruency groups in the Bacteria. Recognition of a succession of others can be expected in the near future, but ultimately these should coalesce to a single grouping that parallels the 16S rRNA tree (except for cases of lateral gene transfer). (ii) The vertical trace of evolution for tryptophan biosynthesis can be deduced. The daunting complexities engendered by paralogy, xenology

  6. Herpes simplex virus type 1 (HSV-1)-derived recombinant vectors for gene transfer and gene therapy.

    PubMed

    Marconi, Peggy; Fraefel, Cornel; Epstein, Alberto L

    2015-01-01

    Herpes simplex virus type 1 (HSV-1 ) is a human pathogen whose lifestyle is based on a long-term dual interaction with the infected host, being able to establish both lytic and latent infections. The virus genome is a 153-kilobase pair (kbp) double-stranded DNA molecule encoding more than 80 genes. The interest of HSV-1 as gene transfer vector stems from its ability to infect many different cell types, both quiescent and proliferating cells, the very high packaging capacity of the virus capsid, the outstanding neurotropic adaptations that this virus has evolved, and the fact that it never integrates into the cellular chromosomes, thus avoiding the risk of insertional mutagenesis. Two types of vectors can be derived from HSV-1, recombinant vectors and amplicon vectors, and different methodologies have been developed to prepare large stocks of each type of vector. This chapter summarizes the approach most commonly used to prepare recombinant HSV-1 vectors through homologous recombination, either in eukaryotic cells or in bacteria.

  7. Interleukin-6 gene transfer reverses body weight gain and fatty liver in obese mice.

    PubMed

    Ma, Yongjie; Gao, Mingming; Sun, Hao; Liu, Dexi

    2015-05-01

    Interleukin-6 (IL-6) is a multifunctional protein and has a major influence on energy metabolism. The current study was designed to assess the therapeutic effect of overexpression of Il-6 gene through gene transfer on high fat diet-induced obese mice. Hydrodynamic delivery of 1 μg pLIVE-IL6 plasmid per mouse into C57BL/6 obese mice resulted in peak level at 10 ng/ml of circulating IL-6 1 day after gene transfer and above 1n g/ml thereafter for a period of 6 weeks. Persistent Il-6 gene expression did not affect food intake but induced a significant reduction in body weight and improved obesity-associated hepatic steatosis. Il-6 gene delivery enhanced thermogenic gene expression and elevated protein levels of phosphorylated STAT3, PGC1α and UCP1 in brown adipose tissue. Il-6 overexpression elevated mRNA levels of lipolysis genes, triggered phosphorylation of STAT3, AMPK, and ACC, and increased expression of genes involved in fatty acid oxidation in skeletal muscle. IL-6 did not affect macrophage infiltration but maintained the M2 macrophage population in adipose tissue. Collectively, these results suggest that overexpression of the Il-6 gene by hydrodynamic gene delivery induces weight loss and alleviates obesity-induced fatty liver and insulin resistance, supporting the notion that gene transfer is a valid approach in managing obesity epidemics.

  8. Vesicular stomatitis virus enables gene transfer and transsynaptic tracing in a wide range of organisms

    PubMed Central

    Mundell, Nathan A.; Beier, Kevin T.; Pan, Y. Albert; Lapan, Sylvain W.; Göz Aytürk, Didem; Berezovskii, Vladimir K.; Wark, Abigail R.; Drokhlyansky, Eugene; Bielecki, Jan; Born, Richard T.; Schier, Alexander F.

    2015-01-01

    Current limitations in technology have prevented an extensive analysis of the connections among neurons, particularly within nonmammalian organisms. We developed a transsynaptic viral tracer originally for use in mice, and then tested its utility in a broader range of organisms. By engineering the vesicular stomatitis virus (VSV) to encode a fluorophore and either the rabies virus glycoprotein (RABV‐G) or its own glycoprotein (VSV‐G), we created viruses that can transsynaptically label neuronal circuits in either the retrograde or anterograde direction, respectively. The vectors were investigated for their utility as polysynaptic tracers of chicken and zebrafish visual pathways. They showed patterns of connectivity consistent with previously characterized visual system connections, and revealed several potentially novel connections. Further, these vectors were shown to infect neurons in several other vertebrates, including Old and New World monkeys, seahorses, axolotls, and Xenopus. They were also shown to infect two invertebrates, Drosophila melanogaster, and the box jellyfish, Tripedalia cystophora, a species previously intractable for gene transfer, although no clear evidence of transsynaptic spread was observed in these species. These vectors provide a starting point for transsynaptic tracing in most vertebrates, and are also excellent candidates for gene transfer in organisms that have been refractory to other methods. J. Comp. Neurol. 523:1639–1663, 2015. © 2015 Wiley Periodicals, Inc. PMID:25688551

  9. Cre-dependent selection yields AAV variants for widespread gene transfer to the adult brain.

    PubMed

    Deverman, Benjamin E; Pravdo, Piers L; Simpson, Bryan P; Kumar, Sripriya Ravindra; Chan, Ken Y; Banerjee, Abhik; Wu, Wei-Li; Yang, Bin; Huber, Nina; Pasca, Sergiu P; Gradinaru, Viviana

    2016-02-01

    Recombinant adeno-associated viruses (rAAVs) are commonly used vehicles for in vivo gene transfer. However, the tropism repertoire of naturally occurring AAVs is limited, prompting a search for novel AAV capsids with desired characteristics. Here we describe a capsid selection method, called Cre recombination-based AAV targeted evolution (CREATE), that enables the development of AAV capsids that more efficiently transduce defined Cre-expressing cell populations in vivo. We use CREATE to generate AAV variants that efficiently and widely transduce the adult mouse central nervous system (CNS) after intravenous injection. One variant, AAV-PHP.B, transfers genes throughout the CNS with an efficiency that is at least 40-fold greater than that of the current standard, AAV9 (refs. 14,15,16,17), and transduces the majority of astrocytes and neurons across multiple CNS regions. In vitro, it transduces human neurons and astrocytes more efficiently than does AAV9, demonstrating the potential of CREATE to produce customized AAV vectors for biomedical applications.

  10. Hydrogen Peroxide-Dependent DNA Release and Transfer of Antibiotic Resistance Genes in Streptococcus gordonii ▿

    PubMed Central

    Itzek, Andreas; Zheng, Lanyan; Chen, Zhiyun; Merritt, Justin; Kreth, Jens

    2011-01-01

    Certain oral streptococci produce H2O2 under aerobic growth conditions to inhibit competing species like Streptococcus mutans. Additionally, H2O2 production causes the release of extracellular DNA (eDNA). eDNA can participate in several important functions: biofilm formation and cell-cell aggregation are supported by eDNA, while eDNA can serve as a nutrient and as an antimicrobial agent by chelating essential cations. eDNA contains DNA fragments of a size that has the potential to transfer genomic information. By using Streptococcus gordonii as a model organism for streptococcal H2O2 production, H2O2-dependent eDNA release was further investigated. Under defined growth conditions, the eDNA release process was shown to be entirely dependent on H2O2. Chromosomal DNA damage seems to be the intrinsic signal for the release, although only actively growing cells were proficient eDNA donors. Interestingly, the process of eDNA production was found to be coupled with the induction of the S. gordonii natural competence system. Consequently, the production of H2O2 triggered the transfer of antibiotic resistance genes. These results suggest that H2O2 is potentially much more than a simple toxic metabolic by-product; rather, its production could serve as an important environmental signal that facilitates species evolution by transfer of genetic information and an increase in the mutation rate. PMID:21984796

  11. Evolutionary transfers of mitochondrial genes to the nucleus in the Populus lineage and coexpression of nuclear and mitochondrial Sdh4 genes.

    PubMed

    Choi, Catherine; Liu, Zhenlan; Adams, Keith L

    2006-01-01

    The transfer of mitochondrial genes to the nucleus is an ongoing evolutionary process in flowering plants. Evolutionarily recent gene transfers provide insights into the evolutionary dynamics of the process and the way in which transferred genes become functional in the nucleus. Genes that are present in the mitochondrion of some angiosperms but have been transferred to the nucleus in the Populus lineage were identified by searches of Populus sequence databases. Sequence analyses and expression experiments were used to characterize the transferred genes. Two succinate dehydrogenase genes and six mitochondrial ribosomal protein genes have been transferred to the nucleus in the Populus lineage and have become expressed. Three transferred genes have gained an N-terminal mitochondrial targeting presequence from other pre-existing genes and two of the transferred genes do not contain an N-terminal targeting presequence. Intact copies of the succinate dehydrogenase gene Sdh4 are present in both the mitochondrion and the nucleus. Both copies of Sdh4 are expressed in multiple organs of two Populus species and RNA editing occurs in the mitochondrial copy. These results provide a genome-wide perspective on mitochondrial genes that were transferred to the nucleus and became expressed, functional genes during the evolutionary history of Populus.

  12. Exploration of horizontal gene transfer between transplastomic tobacco and plant-associated bacteria.

    PubMed

    Demanèche, Sandrine; Monier, Jean-Michel; Dugat-Bony, Eric; Simonet, Pascal

    2011-10-01

    The likelihood of gene transfer from transgenic plants to bacteria is dependent on the transgene copy number and on the presence of homologous sequences for recombination. The large number of chloroplast genomes in a plant cell as well as the prokaryotic origin of the transgene may thus significantly increase the likelihood of gene transfer from transplastomic plants to bacteria. In order to assess the probability of such a transfer, bacterial isolates, screened for their ability to colonize decaying tobacco plant tissue and possessing DNA sequence similarity to the chloroplastic genes accD and rbcL flanking the transgene (aadA), were tested for their ability to take up extracellular DNA (broad host-range pBBR1MCS-3-derived plasmid, transplastomic plant DNA and PCR products containing the genes accD-aadA-rbcL) by natural or electrotransformation. The results showed that among the 16 bacterial isolates tested, six were able to accept foreign DNA and acquire the spectinomycin resistance conferred by the aadA gene on plasmid, but none of them managed to integrate transgenic DNA in their chromosome. Our results provide no indication that the theoretical gene transfer-enhancing properties of transplastomic plants cause horizontal gene transfer at rates above those found in other studies with nuclear transgenes.

  13. Quantitative analysis of recombination between YFP and CFP genes of FRET biosensors introduced by lentiviral or retroviral gene transfer

    PubMed Central

    Komatsubara, Akira T.; Matsuda, Michiyuki; Aoki, Kazuhiro

    2015-01-01

    Biosensors based on the principle of Förster (or fluorescence) resonance energy transfer (FRET) have been developed to visualize spatio-temporal dynamics of signalling molecules in living cells. Many of them adopt a backbone of intramolecular FRET biosensor with a cyan fluorescent protein (CFP) and yellow fluorescent protein (YFP) as donor and acceptor, respectively. However, there remains the difficulty of establishing cells stably expressing FRET biosensors with a YFP and CFP pair by lentiviral or retroviral gene transfer, due to the high incidence of recombination between YFP and CFP genes. To address this, we examined the effects of codon-diversification of YFP on the recombination of FRET biosensors introduced by lentivirus or retrovirus. The YFP gene that was fully codon-optimized to E.coli evaded the recombination in lentiviral or retroviral gene transfer, but the partially codon-diversified YFP did not. Further, the length of spacer between YFP and CFP genes clearly affected recombination efficiency, suggesting that the intramolecular template switching occurred in the reverse-transcription process. The simple mathematical model reproduced the experimental data sufficiently, yielding a recombination rate of 0.002–0.005 per base. Together, these results show that the codon-diversified YFP is a useful tool for expressing FRET biosensors by lentiviral or retroviral gene transfer. PMID:26290434

  14. Quantitative analysis of recombination between YFP and CFP genes of FRET biosensors introduced by lentiviral or retroviral gene transfer.

    PubMed

    Komatsubara, Akira T; Matsuda, Michiyuki; Aoki, Kazuhiro

    2015-08-20

    Biosensors based on the principle of Förster (or fluorescence) resonance energy transfer (FRET) have been developed to visualize spatio-temporal dynamics of signalling molecules in living cells. Many of them adopt a backbone of intramolecular FRET biosensor with a cyan fluorescent protein (CFP) and yellow fluorescent protein (YFP) as donor and acceptor, respectively. However, there remains the difficulty of establishing cells stably expressing FRET biosensors with a YFP and CFP pair by lentiviral or retroviral gene transfer, due to the high incidence of recombination between YFP and CFP genes. To address this, we examined the effects of codon-diversification of YFP on the recombination of FRET biosensors introduced by lentivirus or retrovirus. The YFP gene that was fully codon-optimized to E.coli evaded the recombination in lentiviral or retroviral gene transfer, but the partially codon-diversified YFP did not. Further, the length of spacer between YFP and CFP genes clearly affected recombination efficiency, suggesting that the intramolecular template switching occurred in the reverse-transcription process. The simple mathematical model reproduced the experimental data sufficiently, yielding a recombination rate of 0.002-0.005 per base. Together, these results show that the codon-diversified YFP is a useful tool for expressing FRET biosensors by lentiviral or retroviral gene transfer.

  15. Horizontal transfers of transposable elements in eukaryotes: The flying genes.

    PubMed

    Panaud, Olivier

    2016-01-01

    Transposable elements (TEs) are the major components of eukaryotic genomes. Their propensity to densely populate and in some cases invade the genomes of plants and animals is in contradiction with the fact that transposition is strictly controlled by several molecular pathways acting at either transcriptional or post-transcriptional levels. Horizontal transfers, defined as the transmission of genetic material between sexually isolated species, have long been considered as rare phenomena. Here, we show that the horizontal transfers of transposable elements (HTTs) are very frequent in ecosystems. The exact mechanisms of such transfers are not well understood, but species involved in close biotic interactions, like parasitism, show a propensity to exchange genetic material horizontally. We propose that HTTs allow TEs to escape the silencing machinery of their host genome and may therefore be an important mechanism for their survival and their dissemination in eukaryotes.

  16. Cooperative heat transfer and ground coupled storage system

    DOEpatents

    Metz, Philip D.

    1982-01-01

    A cooperative heat transfer and ground coupled storage system wherein collected solar heat energy is ground stored and permitted to radiate into the adjacent ground for storage therein over an extended period of time when such heat energy is seasonally maximally available. Thereafter, when said heat energy is seasonally minimally available and has propagated through the adjacent ground a substantial distance, the stored heat energy may be retrieved by a circumferentially arranged heat transfer means having a high rate of heat transfer.

  17. Cooperative heat transfer and ground coupled storage system

    DOEpatents

    Metz, P.D.

    A cooperative heat transfer and ground coupled storage system wherein collected solar heat energy is ground stored and permitted to radiate into the adjacent ground for storage therein over an extended period of time when such heat energy is seasonally maximally available. Thereafter, when said heat energy is seasonally minimally available and has propagated through the adjacent ground a substantial distance, the stored heat energy may be retrieved by a circumferentially arranged heat transfer means having a high rate of heat transfer.

  18. General Relativistic Radiative Transfer: Applications to Black-Hole Systems

    NASA Technical Reports Server (NTRS)

    Wu, Kinwah; Fuerst, Steven V.; Mizuno, Yosuke; Nishikawa, Ken-Ichi; Branduardi-Raymont, Graziella; Lee, Khee-Gan

    2007-01-01

    We present general relativistic radiation transfer formulations which include opacity effects due to absorption, emission and scattering explicitly. We consider a moment expansions for the transfer in the presence of scattering. The formulation is applied to calculation emissions from accretion and outflows in black-hole systems. Cases with thin accretion disks and accretion tori are considered. Effects, such as emission anisotropy, non-stationary flows and geometrical self-occultation are investigated. Polarisation transfer in curved space-time is discussed qualitatively.

  19. Resurrection of an ancestral gene: functional and evolutionary analyses of the Ngrol genes transferred from Agrobacterium to Nicotiana.

    PubMed

    Aoki, Seishiro

    2004-08-01

    The Ng rol genes, which have high similarity in sequence to the rol genes of Agrobacterium rhizogenes, are present in the genome of untransformed plants of Nicotiana glauca. It is thought that bacterial infection resulted in the transfer of the Ng rol genes to plants early in the evolution of the genus Nicotiana, since several species in this genus contain rol-like sequences but others do not. Plants transformed with the bacterial rol genes exhibit various developmental and morphological changes. The presence of rol-like sequences in plant genomes is therefore thought to have contributed to the evolution of Nicotiana species. This paper focuses on studies of the Ng rol genes in present-day plants and during the evolution of the genus Nicotiana. The functional sequences of several Ng rol genes may have been conserved after their ancient introduction from a bacterium to the plant. Resurrection of an ancestral function of one of the Ng rol genes, as examined by physiological and evolutionary analyses, is also described. The origin of the Ng rol genes is then considered, based on results of molecular phylogenetic analyses. The effects of the horizontal transfer of the Ng rol genes and mutations in the genes are discussed on the plants of the genus Nicotiana during evolution.

  20. Phylogenetic analysis of the incidence of lux gene horizontal transfer in Vibrionaceae.

    PubMed

    Urbanczyk, Henryk; Ast, Jennifer C; Kaeding, Allison J; Oliver, James D; Dunlap, Paul V

    2008-05-01

    Horizontal gene transfer (HGT) is thought to occur frequently in bacteria in nature and to play an important role in bacterial evolution, contributing to the formation of new species. To gain insight into the frequency of HGT in Vibrionaceae and its possible impact on speciation, we assessed the incidence of interspecies transfer of the lux genes (luxCDABEG), which encode proteins involved in luminescence, a distinctive phenotype. Three hundred three luminous strains, most of which were recently isolated from nature and which represent 11 Aliivibrio, Photobacterium, and Vibrio species, were screened for incongruence of phylogenies based on a representative housekeeping gene (gyrB or pyrH) and a representative lux gene (luxA). Strains exhibiting incongruence were then subjected to detailed phylogenetic analysis of horizontal transfer by using multiple housekeeping genes (gyrB, recA, and pyrH) and multiple lux genes (luxCDABEG). In nearly all cases, housekeeping gene and lux gene phylogenies were congruent, and there was no instance in which the lux genes of one luminous species had replaced the lux genes of another luminous species. Therefore, the lux genes are predominantly vertically inherited in Vibrionaceae. The few exceptions to this pattern of congruence were as follows: (i) the lux genes of the only known luminous strain of Vibrio vulnificus, VVL1 (ATCC 43382), were evolutionarily closely related to the lux genes of Vibrio harveyi; (ii) the lux genes of two luminous strains of Vibrio chagasii, 21N-12 and SB-52, were closely related to those of V. harveyi and Vibrio splendidus, respectively; (iii) the lux genes of a luminous strain of Photobacterium damselae, BT-6, were closely related to the lux genes of the lux-rib(2) operon of Photobacterium leiognathi; and (iv) a strain of the luminous bacterium Photobacterium mandapamensis was found to be merodiploid for the lux genes, and the second set of lux genes was closely related to the lux genes of the lux-rib(2

  1. Phylogenetic Analysis of the Incidence of lux Gene Horizontal Transfer in Vibrionaceae▿ †

    PubMed Central

    Urbanczyk, Henryk; Ast, Jennifer C.; Kaeding, Allison J.; Oliver, James D.; Dunlap, Paul V.

    2008-01-01

    Horizontal gene transfer (HGT) is thought to occur frequently in bacteria in nature and to play an important role in bacterial evolution, contributing to the formation of new species. To gain insight into the frequency of HGT in Vibrionaceae and its possible impact on speciation, we assessed the incidence of interspecies transfer of the lux genes (luxCDABEG), which encode proteins involved in luminescence, a distinctive phenotype. Three hundred three luminous strains, most of which were recently isolated from nature and which represent 11 Aliivibrio, Photobacterium, and Vibrio species, were screened for incongruence of phylogenies based on a representative housekeeping gene (gyrB or pyrH) and a representative lux gene (luxA). Strains exhibiting incongruence were then subjected to detailed phylogenetic analysis of horizontal transfer by using multiple housekeeping genes (gyrB, recA, and pyrH) and multiple lux genes (luxCDABEG). In nearly all cases, housekeeping gene and lux gene phylogenies were congruent, and there was no instance in which the lux genes of one luminous species had replaced the lux genes of another luminous species. Therefore, the lux genes are predominantly vertically inherited in Vibrionaceae. The few exceptions to this pattern of congruence were as follows: (i) the lux genes of the only known luminous strain of Vibrio vulnificus, VVL1 (ATCC 43382), were evolutionarily closely related to the lux genes of Vibrio harveyi; (ii) the lux genes of two luminous strains of Vibrio chagasii, 21N-12 and SB-52, were closely related to those of V. harveyi and Vibrio splendidus, respectively; (iii) the lux genes of a luminous strain of Photobacterium damselae, BT-6, were closely related to the lux genes of the lux-rib2 operon of Photobacterium leiognathi; and (iv) a strain of the luminous bacterium Photobacterium mandapamensis was found to be merodiploid for the lux genes, and the second set of lux genes was closely related to the lux genes of the lux-rib2

  2. Comparative genomics of Neisseria meningitidis: core genome, islands of horizontal transfer and pathogen-specific genes.

    PubMed

    Dunning Hotopp, Julie C; Grifantini, Renata; Kumar, Nikhil; Tzeng, Yih Ling; Fouts, Derrick; Frigimelica, Elisabetta; Draghi, Monia; Giuliani, Marzia Monica; Rappuoli, Rino; Stephens, David S; Grandi, Guido; Tettelin, Hervé

    2006-12-01

    To better understand Neisseria meningitidis genomes and virulence, microarray comparative genome hybridization (mCGH) data were collected from one Neisseria cinerea, two Neisseria lactamica, two Neisseria gonorrhoeae and 48 Neisseria meningitidis isolates. For N. meningitidis, these isolates are from diverse clonal complexes, invasive and carriage strains, and all major serogroups. The microarray platform represented N. meningitidis strains MC58, Z2491 and FAM18, and N. gonorrhoeae FA1090. By comparing hybridization data to genome sequences, the core N. meningitidis genome and insertions/deletions (e.g. capsule locus, type I secretion system) related to pathogenicity were identified, including further characterization of the capsule locus, bioinformatics analysis of a type I secretion system, and identification of some metabolic pathways associated with intracellular survival in pathogens. Hybridization data clustered meningococcal isolates from similar clonal complexes that were distinguished by the differential presence of six distinct islands of horizontal transfer. Several of these islands contained prophage or other mobile elements, including a novel prophage and a transposon carrying portions of a type I secretion system. Acquisition of some genetic islands appears to have occurred in multiple lineages, including transfer between N. lactamica and N. meningitidis. However, island acquisition occurs infrequently, such that the genomic-level relationship is not obscured within clonal complexes. The N. meningitidis genome is characterized by the horizontal acquisition of multiple genetic islands; the study of these islands reveals important sets of genes varying between isolates and likely to be related to pathogenicity.

  3. Interleukin 10 gene transfer prevents experimental colitis in rats

    PubMed Central

    Barbara, G; Xing, Z; Hogaboam, C; Gauldie, J; Collins, S

    2000-01-01

    BACKGROUND—The development of colitis in interleukin 10 (IL-10) deficient mice, together with the known anti-inflammatory and immunomodulatory properties of this cytokine have prompted consideration of IL-10 as a treatment for inflammatory bowel disease (IBD). However, studies using hrIL-10 in IBD models have yielded inconsistent results.
AIMS—To examine the therapeutic potential of overexpressing the IL-10 gene before and after the induction of experimental colitis in rats.
METHODS—Gene transfer was achieved by intraperitoneal injection of non-replicating human type 5 adenovirus bearing the IL-10 gene, either 24 hours before or one hour after intrarectal administration of dinitrobenzene sulphonic acid in rats. Colonic damage and inflammation was assessed macroscopically and by measuring myeloperoxidase activity and leukotriene B4 concentrations.
RESULTS—Gene transfer increased IL-10 protein in serum for up to six days. IL-10 gene transfer prior to colitis improved colitis macroscopically and histologically, and significantly reduced colonic myeloperoxidase activity and leukotriene B4 concentrations. In contrast, IL-10 gene transfer after the onset of colitis had no beneficial effect.
CONCLUSIONS—Gene therapy using an adenovirus-IL-10 construct was successful in preventing but not in reversing experimental colitis in the rat.


Keywords: gene therapy; colitis; interleukin 10; adenovirus vector; leukotrienes; inflammatory bowel disease; maintenance therapy PMID:10673295

  4. Multilevel populations and the evolution of antibiotic resistance through horizontal gene transfer.

    PubMed

    Andam, Cheryl P; Fournier, Gregory P; Gogarten, Johann Peter

    2011-09-01

    Horizontal gene transfer (HGT) can create diversity in the genetic repertoire of a lineage. Successful gene transfer likely occurs more frequently between more closely related organisms, leading to the formation of higher-level exchange groups that in some respects are comparable to single-species populations. Genes that appear fixed in a single species can be replaced through distant homologs or iso-functional analogs acquired through HGT. These genes may originate from other species or they may be acquired by an individual strain from the species pan-genome. Because of their similarity to alleles in a population, we label these gene variants that are exchanged between related species as homeoalleles. In a case study, we show that biased gene transfer plays an important role in the evolution of aminoacyl-tRNA synthetases (aaRS). Many microorganisms make use of these genes against naturally occurring antibiotics. We suggest that the resistance against naturally occurring antibiotics is the likely driving force behind the frequent switching between divergent aaRS types and the reason for the maintenance of these homeoalleles in higher-level exchange groups. Resistance to naturally occurring antibiotics may lead to the maintenance of different types of aminoacyl-tRNA synthetases in Bacteria through gene transfer.

  5. Studies on the transfer techniques of three maize genes.

    PubMed

    Wang, G; Zhang, H; Ding, Q; Xie, Y; Dai, J

    1996-01-01

    Maize transformation has been carried out through microprojectile bombardment, ultrasonication in a DNA buffer, and ovary-injection with a self-made microinjector. The plasmid pB48.415, which carries a 3'-truncated Bt-toxin protein gene and a hygromycin phosphotransferase (hpt) gene, was used in the transformation. Transgenic maize plants were obtained from immature embryos and embryogenic calli bombarded with a particle gun, embryogenic calli ultrasonicated under different conditions of ovaries injected 10-20 hours after pollination. The results of Dot blotting and Southern blotting analyses proved the integration of the Bt gene into maize genome.

  6. Extensive recombination events and horizontal gene transfer shaped the Legionella pneumophila genomes

    PubMed Central

    2011-01-01

    Background Legionella pneumophila is an intracellular pathogen of environmental protozoa. When humans inhale contaminated aerosols this bacterium may cause a severe pneumonia called Legionnaires' disease. Despite the abundance of dozens of Legionella species in aquatic reservoirs, the vast majority of human disease is caused by a single serogroup (Sg) of a single species, namely L. pneumophila Sg1. To get further insights into genome dynamics and evolution of Sg1 strains, we sequenced strains Lorraine and HL 0604 1035 (Sg1) and compared them to the available sequences of Sg1 strains Paris, Lens, Corby and Philadelphia, resulting in a comprehensive multigenome analysis. Results We show that L. pneumophila Sg1 has a highly conserved and syntenic core genome that comprises the many eukaryotic like proteins and a conserved repertoire of over 200 Dot/Icm type IV secreted substrates. However, recombination events and horizontal gene transfer are frequent. In particular the analyses of the distribution of nucleotide polymorphisms suggests that large chromosomal fragments of over 200 kbs are exchanged between L. pneumophila strains and contribute to the genome dynamics in the natural population. The many secretion systems present might be implicated in exchange of these fragments by conjugal transfer. Plasmids also play a role in genome diversification and are exchanged among strains and circulate between different Legionella species. Conclusion Horizontal gene transfer among bacteria and from eukaryotes to L. pneumophila as well as recombination between strains allows different clones to evolve into predominant disease clones and others to replace them subsequently within relatively short periods of time. PMID:22044686

  7. Horizontal Gene Transfers from Bacteria to Entamoeba Complex: A Strategy for Dating Events along Species Divergence

    PubMed Central

    Romero, Miguel; Ximenez, Cecilia

    2016-01-01

    Horizontal gene transfer has proved to be relevant in eukaryotic evolution, as it has been found more often than expected and related to adaptation to certain niches. A relatively large list of laterally transferred genes has been proposed and evaluated for the parasite Entamoeba histolytica. The goals of this work were to elucidate the importance of lateral gene transfer along the evolutionary history of some members of the genus Entamoeba, through identifying donor groups and estimating the divergence time of some of these events. In order to estimate the divergence time of some of the horizontal gene transfer events, the dating of some Entamoeba species was necessary, following an indirect dating strategy based on the fossil record of plausible hosts. The divergence between E. histolytica and E. nuttallii probably occurred 5.93 million years ago (Mya); this lineage diverged from E. dispar 9.97 Mya, while the ancestor of the latter separated from E. invadens 68.18 Mya. We estimated times for 22 transferences; the most recent occurred 31.45 Mya and the oldest 253.59 Mya. Indeed, the acquisition of genes through lateral transfer may have triggered a period of adaptive radiation, thus playing a major role in the evolution of the Entamoeba genus. PMID:27239333

  8. Cre Recombinase Gene Transfer In Vitro and Detection of loxP-Dependent Recombination.

    PubMed

    Ohtsubo, Kazuaki; Marth, Jamey D

    2007-07-01

    INTRODUCTIONAltering the genome of intact cells and organisms by site-specific DNA recombination has become an important gene-transfer methodology. The inclusion of exogenous recombinase target sequences within transferred DNA segments allows subsequent modifications to previously altered genomic structure that increase the utility of gene transfer and enhance experimental design. In this protocol, correctly targeted mouse embryonic stem (ES) cell clones bearing the F[tkneo] allele (containing several loxP sites) are subjected to in vitro Cre gene transfer to generate ES cell subclones bearing either Type 1 (Δ) or Type 2 (F) alleles. Type 2 ES cells are used to generate chimeric mice that are then crossed to germ-line Cre-expressing mice, such as ZP3-Cre transgenic mates. The additional time needed to breed the mice (~2-3 mo) is typically less troublesome than the cost and effort of maintaining multiple clone-derived lines of mice.

  9. Diversity, evolution, and horizontal gene transfer (HGT) in soda lakes

    NASA Astrophysics Data System (ADS)

    Pinkart, Holly C.; Storrie-Lombardi, Michael C.

    2007-09-01

    Soap Lake is a hypersaline, alkaline lake in Central Washington State (USA). For the past five years the lake has been the site of an NSF Microbial Observatory project devoted to identifying critical geochemical and microbial characteristics of the monimolimnion sediment and water column, and has demonstrated rich multispecies communities occupy all areas of the lake. Soap Lake and similar soda lakes are subject to repeated transient periods of extreme evaporation characterized by significant repetitive alterations in salinity, pH, and total water volume, yet maintain high genetic and metabolic diversity. It has been argued that this repetitive cycle for salinity, alkalinity, and sulfur concentration has been a major driver for prokaryote evolution and diversity. The rapidity of wet-dry cycling places special demands on genome evolution, requirements that are beyond the relatively conservative eukaryotic evolutionary strategy of serial alteration of existing gene sequences in a relatively stable genome. Although HGT is most likely responsible for adding a significant amount of noise to the genetic record, analysis of HGT activity can also provide us with a much-needed probe for exploration of prokaryotic genome evolution and the origin of diversity. Packaging of genetic information within the protective protein capsid of a bacteriophage would seem preferable to exposing naked DNA to the highly alkaline conditions in the lake. In this study, we present preliminary data demonstrating the presence of a diverse group of phage integrases in Soap Lake. Integrase is the viral enzyme responsible for the insertion of phage DNA into the bacterial host's chromosome. The presence of the integrase sequence in bacterial chromosomes is evidence of lysogeny, and the diversity of integrase sequences reported here suggests a wide variety of temperate phage exist in this system, and are especially active in transition zones.

  10. Development of an Autologous Macrophage-based Adoptive Gene Transfer Strategy to Treat Posttraumatic Osteoarthritis (PTOA) and Osteoarithritis (OA)

    DTIC Science & Technology

    2016-05-01

    AWARD NUMBER: W81XWH-13-1-0228 TITLE: Development of an Autologous Macrophage-based Adoptive Gene Transfer Strategy to Treat Posttraumatic...Final 3. DATES COVERED 1 Sep 2013 - 28 Feb 2016 4. TITLE AND SUBTITLE Development of an Autologous Macrophage-based Adoptive Gene Transfer Strategy to...autologous macrophage-based adoptive gene transfer strategy can effectively deliver and confine expression of an anti-catabolic gene (IL-1ra or IL-1β

  11. Evaluation of Vascular Delivery Methodologies to Enhance rAAV6-mediated Gene Transfer to Canine Striated Musculature

    PubMed Central

    Gregorevic, Paul; Schultz, Brian R; Allen, James M; Halldorson, Jeffrey B; Blankinship, Michael J; Meznarich, Norman A; Kuhr, Christian S; Doremus, Caitlin; Finn, Eric; Liggitt, Denny; Chamberlain, Jeffrey S

    2009-01-01

    A growing body of research supports the development of recombinant adeno-associated viral (rAAV) vectors for delivery of gene expression cassettes to striated musculature as a method of treating severe neuromuscular conditions. However, it is unclear whether delivery protocols that achieve extensive gene transfer in mice can be adapted to produce similarly extensive gene transfer in larger mammals and ultimately patients. Consequently, we sought to investigate methodological modifications that would facilitate rAAV-mediated gene transfer to the striated musculature of canines. A simple procedure incorporating acute (i) occlusion of limb blood flow, (ii) exsanguination via compression bandage, and (iii) vector “dwell” time of <20 minutes, markedly enhanced the transduction of limb muscles, compared with a simple bolus limb infusion of vector. A complementary method whereby vector was infused into the jugular vein led to efficient transduction of cardiomyocytes and to a lesser degree the diaphragm. Together these methods can be used to achieve transgene expression in heart, diaphragm, and limb muscles of juvenile dogs using rAAV6 vectors. These results establish that rAAV-mediated gene delivery is a viable approach to achieving systemic transduction of striated musculature in mammals approaching the dimensions of newborn humans. PMID:19471246

  12. Linear topology confers in vivo gene transfer activity to polyethylenimines.

    PubMed

    Brissault, B; Leborgne, C; Guis, C; Danos, O; Cheradame, H; Kichler, A

    2006-01-01

    Although polyethylenimines (PEIs) are frequently used transfection agents, it is still unclear which of their properties are required for efficient gene delivery. This is even more striking when working in vivo since some PEIs are able to generate significant gene expression, whereas others are not. To facilitate a rational development of compounds with improved transfection activities, studies aimed at identifying the properties involved in the transfection process seem indispensable. In the present work, we investigated how transfection with linear PEI of 22 kDa allows for high reporter gene expression in lungs after intravenous injection, whereas the branched PEI of 25 kDa does not. To this end, we synthesized L-PEI derivatives that are intermediates between linear and branched PEIs. Our results show that the topology plays a crucial role in obtaining in vivo reporter gene expression, whereas the content of primary, secondary, and tertiary amines is only of minor importance.

  13. Horizontal Gene Transfer and the Evolution of Bacterial and Archaeal Population Structure

    PubMed Central

    Alm, Eric J.; Hanage, William P.

    2013-01-01

    Many bacterial and archaeal lineages have a history of extensive and ongoing horizontal gene transfer and loss, as evidenced by the large differences in genome content even among otherwise closely related isolates. How ecologically cohesive populations might evolve and be maintained under such conditions of rapid gene turnover has remained controversial. Here we synthesize recent literature demonstrating the importance of habitat and niche in structuring horizontal gene transfer. This leads to a model of ecological speciation via gradual genetic isolation triggered by differential habitat association of nascent populations. Further, we hypothesize that subpopulations can evolve through local gene exchange networks by tapping into a gene pool that is adaptive towards local, continuously changing organismic interactions and is, to a large degree, responsible for the observed rapid gene turnover. Overall, these insights help explain how bacteria and archaea form populations that display both ecological cohesion and high genomic diversity. PMID:23332119

  14. Introduction to the Subjective Transfer Function Approach to Analyzing Systems.

    DTIC Science & Technology

    1984-07-01

    STANDARtDS- 1963-A R-3021-AF Intoduction to the Subjective Transfer Function Approach to Analyzing Systems 00 • CO Lf. Clairice T. Veit, Monti Callero ...34Prepared for the United States Air Force." Bibliography: p. • "R-3021-AF." 1. Subjective transfer function method. 2. System analysis. I. Callero , Monti...to Analyzing Systems T, Clairice T. Veit, Monti Callero , Barbara J. Rose July 1984 A Project AIR FORCE report prepared for the - United States Air

  15. Split vector systems for ultra-targeted gene delivery: a contrivance to achieve ethical assurance of somatic gene therapy in vivo.

    PubMed

    Tolmachov, Oleg E

    2014-08-01

    Tightly controlled spatial localisation of therapeutic gene delivery is essential to maximize the benefits of somatic gene therapy in vivo and to reduce its undesired effects on the 'bystander' cell populations, most importantly germline cells. Indeed, complete ethical assurance of somatic gene therapy can only be achieved with ultra-targeted gene delivery, which excludes the risk of inadvertent germline gene transfer. Thus, it is desired to supplement existing strategies of physical focusing and biological (cell-specific) targeting of gene delivery with an additional principle for the rigid control over spread of gene transfer within the body. In this paper I advance the concept of 'combinatorial' targeting of therapeutic gene transfer in vivo. I hypothesize that it is possible to engineer complex gene delivery vector systems consisting of several components, each one of them capable of independent spread within the human body but incapable of independent facilitation of gene transfer. As the gene delivery augmented by such split vector systems would be reliant on the simultaneous availability of all the vector system components at a predetermined body site, it is envisaged that higher order reaction kinetics required for the assembly of the functional gene transfer configuration would sharpen spatial localisation of gene transfer via curtailing the blurring effect of the vector spread within the body. A particular implementation of such split vector system could be obtained through supplementing a viral therapeutic gene vector with a separate auxiliary vector carrying a non-integrative and non-replicative form of a gene (e.g., mRNA) coding for a cellular receptor of the therapeutic vector component. Gene-transfer-enabling components of the vector system, which would be delivered separately from the vector component loaded with the therapeutic gene cargo, could also be cell-membrane-insertion-proficient receptors, elements of artificial transmembrane channels

  16. Targeted gene transfer into rat facial muscles by nanosecond pulsed laser-induced stress waves.

    PubMed

    Kurita, Akihiro; Matsunobu, Takeshi; Satoh, Yasushi; Ando, Takahiro; Sato, Shunichi; Obara, Minoru; Shiotani, Akihiro

    2011-09-01

    We investigate the feasibility of using nanosecond pulsed laser-induced stress waves (LISWs) for gene transfer into rat facial muscles. LISWs are generated by irradiating a black natural rubber disk placed on the target tissue with nanosecond pulsed laser light from the second harmonics (532 nm) of a Q-switched Nd:YAG laser, which is widely used in head and neck surgery and proven to be safe. After injection of plasmid deoxyribose nucleic acid (DNA) coding for Lac Z into rat facial muscles, pulsed laser is used to irradiate the laser target on the skin surface without incision or exposure of muscles. Lac Z expression is detected by X-gal staining of excised rat facial skin and muscles. Strong Lac Z expression is observed seven days after gene transfer, and sustained for up to 14 days. Gene transfer is achieved in facial muscles several millimeters deep from the surface. Gene expression is localized to the tissue exposed to LISWs. No tissue damage from LISWs is observed. LISW is a promising nonviral target gene transfer method because of its high spatial controllability, easy applicability, and minimal invasiveness. Gene transfer using LISW to produce therapeutic proteins such as growth factors could be used to treat nerve injury and paralysis.

  17. Targeted gene transfer into rat facial muscles by nanosecond pulsed laser-induced stress waves

    NASA Astrophysics Data System (ADS)

    Kurita, Akihiro; Matsunobu, Takeshi; Satoh, Yasushi; Ando, Takahiro; Sato, Shunichi; Obara, Minoru; Shiotani, Akihiro

    2011-09-01

    We investigate the feasibility of using nanosecond pulsed laser-induced stress waves (LISWs) for gene transfer into rat facial muscles. LISWs are generated by irradiating a black natural rubber disk placed on the target tissue with nanosecond pulsed laser light from the second harmonics (532 nm) of a Q-switched Nd:YAG laser, which is widely used in head and neck surgery and proven to be safe. After injection of plasmid deoxyribose nucleic acid (DNA) coding for Lac Z into rat facial muscles, pulsed laser is used to irradiate the laser target on the skin surface without incision or exposure of muscles. Lac Z expression is detected by X-gal staining of excised rat facial skin and muscles. Strong Lac Z expression is observed seven days after gene transfer, and sustained for up to 14 days. Gene transfer is achieved in facial muscles several millimeters deep from the surface. Gene expression is localized to the tissue exposed to LISWs. No tissue damage from LISWs is observed. LISW is a promising nonviral target gene transfer method because of its high spatial controllability, easy applicability, and minimal invasiveness. Gene transfer using LISW to produce therapeutic proteins such as growth factors could be used to treat nerve injury and paralysis.

  18. Complexity of genetic sequences modified by horizontal gene transfer and degraded-DNA uptake

    NASA Astrophysics Data System (ADS)

    Tremberger, George; Dehipawala, S.; Nguyen, A.; Cheung, E.; Sullivan, R.; Holden, T.; Lieberman, D.; Cheung, T.

    2015-09-01

    Horizontal gene transfer has been a major vehicle for efficient transfer of genetic materials among living species and could be one of the sources for noncoding DNA incorporation into a genome. Our previous study of lnc- RNA sequence complexity in terms of fractal dimension and information entropy shows a tight regulation among the studied genes in numerous diseases. The role of sequence complexity in horizontal transferred genes was investigated with Mealybug in symbiotic relation with a 139K genome microbe and Deinococcus radiodurans as examples. The fractal dimension and entropy showed correlation R-sq of 0.82 (N = 6) for the studied Deinococcus radiodurans sequences. For comparison the Deinococcus radiodurans oxidative stress tolerant catalase and superoxide dismutase genes under extracellular dGMP growth condition showed R-sq ~ 0.42 (N = 6); and the studied arsenate reductase horizontal transferred genes for toxicity survival in several microorganisms showed no correlation. Simulation results showed that R-sq < 0.4 would be improbable at less than one percent chance, suggestive of additional selection pressure when compared to the R-sq ~ 0.29 (N = 21) in the studied transferred genes in Mealybug. The mild correlation of R-sq ~ 0.5 for fractal dimension versus transcription level in the studied Deinococcus radiodurans sequences upon extracellular dGMP growth condition would suggest that lower fractal dimension with less electron density fluctuation favors higher transcription level.

  19. Gene transfers shaped the evolution of de novo NAD+ biosynthesis in eukaryotes.

    PubMed

    Ternes, Chad M; Schönknecht, Gerald

    2014-09-01

    NAD(+) is an essential molecule for life, present in each living cell. It can function as an electron carrier or cofactor in redox biochemistry and energetics, and serves as substrate to generate the secondary messenger cyclic ADP ribose and nicotinic acid adenine dinucleotide phosphate. Although de novo NAD(+) biosynthesis is essential, different metabolic pathways exist in different eukaryotic clades. The kynurenine pathway starting with tryptophan was most likely present in the last common ancestor of all eukaryotes, and is active in fungi and animals. The aspartate pathway, detected in most photosynthetic eukaryotes, was probably acquired from the cyanobacterial endosymbiont that gave rise to chloroplasts. An evolutionary analysis of enzymes catalyzing de novo NAD(+) biosynthesis resulted in evolutionary trees incongruent with established organismal phylogeny, indicating numerous gene transfers. Endosymbiotic gene transfers probably introduced the aspartate pathway into eukaryotes and may have distributed it among different photosynthetic clades. In addition, several horizontal gene transfers substituted eukaryotic genes with bacterial orthologs. Although horizontal gene transfer is accepted as a key mechanism in prokaryotic evolution, it is supposed to be rare in eukaryotic evolution. The essential metabolic pathway of de novo NAD(+) biosynthesis in eukaryotes was shaped by numerous gene transfers.

  20. Provincial Postsecondary Systems and Arrangements for Credit Transfer.

    ERIC Educational Resources Information Center

    British Columbia Council on Admissions and Transfer, Vancouver.

    This document describes the structure of the postsecondary education system, the mechanisms to facilitate transfer and articulation, and the major instruments used to facilitate transfer and articulation for postsecondary institutions in the Canadian Provinces. According to this document, the British Columbia postsecondary educational school…

  1. Radiation heat transfer shapefactors for combustion systems

    NASA Technical Reports Server (NTRS)

    Emery, A. F.; Johansson, O.; Abrous, A.

    1987-01-01

    The computation of radiation heat transfer through absorbing media is commonly done through the zoning method which relies upon values of the geometric mean transmittance and absorptance. The computation of these values is difficult and expensive, particularly if many spectral bands are used. This paper describes the extension of a scan line algorithm, based upon surface-surface radiation, to the computation of surface-gas and gas-gas radiation transmittances.

  2. Transfer Learning in Integrated Cognitive Systems

    DTIC Science & Technology

    2010-09-01

    with the research on MLNs lead by Pedro Domingos at the University of Washington (UW). All of the software developed was integrated with UW’s Alchemy ...learning methods. We refer to the structure-learning method developed by Kok and Domingos (2005) as Alchemy . We ran Alchemy “from scratch” without...transfer learning ( Alchemy Scratch); as well as using it to revise the MLN learned in the source domain and then automatically mapped to the target

  3. The agricultural antibiotic carbadox induces phage-mediated gene transfer in Salmonella

    PubMed Central

    Bearson, Bradley L.; Allen, Heather K.; Brunelle, Brian W.; Lee, In Soo; Casjens, Sherwood R.; Stanton, Thaddeus B.

    2013-01-01

    Antibiotics are used for disease therapeutic or preventative effects in humans and animals, as well as for enhanced feed conversion efficiency in livestock. Antibiotics can also cause undesirable effects in microbial populations, including selection for antibiotic resistance, enhanced pathogen invasion, and stimulation of horizontal gene transfer. Carbadox is a veterinary antibiotic used in the US during the starter phase of swine production for improved feed efficiency and control of swine dysentery and bacterial swine enteritis. Carbadox has been shown in vitro to induce phage-encoded Shiga toxin in Shiga toxin-producing Escherichia coli (STEC) and a phage-like element transferring antibiotic resistance genes in Brachyspira hyodysenteriae, but the effect of carbadox on prophages in other bacteria is unknown. This study examined carbadox exposure on prophage induction and genetic transfer in Salmonella enterica serovar Typhimurium, a human foodborne pathogen that frequently colonizes swine without causing disease. S. Typhimurium LT2 exposed to carbadox induced prophage production, resulting in bacterial cell lysis and release of virions that were visible by electron microscopy. Carbadox induction of phage-mediated gene transfer was confirmed by monitoring the transduction of a sodCIII::neo cassette in the Fels-1 prophage from LT2 to a recipient Salmonella strain. Furthermore, carbadox frequently induced generalized transducing phages in multidrug-resistant phage type DT104 and DT120 isolates, resulting in the transfer of chromosomal and plasmid DNA that included antibiotic resistance genes. Our research indicates that exposure of Salmonella to carbadox induces prophages that can transfer virulence and antibiotic resistance genes to susceptible bacterial hosts. Carbadox-induced, phage-mediated gene transfer could serve as a contributing factor in bacterial evolution during animal production, with prophages being a reservoir for bacterial fitness genes in the

  4. An rf communications system for the West Valley transfer cart

    SciTech Connect

    Crutcher, R.I.; Moore, M.R.

    1993-01-01

    A prototype radio frequency communications system for digital data was designed and built by Oak Ridge National Laboratory for use in controlling the vitrification facility transfer cart at the West Valley Nuclear Services facility in New York. The communications system provides bidirectional wireless data transfer between the operator control station and the material transfer cart. The system was designed to operate in radiation fields of 10[sup 4] R/h while withstanding a total integrated dose of 10[sup 7] R of gamma radiation. Implementation of antenna spatial diversity, automatic gain control, and spectral processing improves operation in the reflective environment of the metal-lined reprocessing cells.

  5. An rf communications system for the West Valley transfer cart

    SciTech Connect

    Crutcher, R.I.; Moore, M.R.

    1993-05-01

    A prototype radio frequency communications system for digital data was designed and built by Oak Ridge National Laboratory for use in controlling the vitrification facility transfer cart at the West Valley Nuclear Services facility in New York. The communications system provides bidirectional wireless data transfer between the operator control station and the material transfer cart. The system was designed to operate in radiation fields of 10{sup 4} R/h while withstanding a total integrated dose of 10{sup 7} R of gamma radiation. Implementation of antenna spatial diversity, automatic gain control, and spectral processing improves operation in the reflective environment of the metal-lined reprocessing cells.

  6. Horizontal Gene Transfer of Phytochelatin Synthases from Bacteria to Extremophilic Green Algae.

    PubMed

    Olsson, Sanna; Penacho, Vanessa; Puente-Sánchez, Fernando; Díaz, Silvia; Gonzalez-Pastor, José Eduardo; Aguilera, Angeles

    2017-01-01

    Transcriptomic sequencing together with bioinformatic analyses and an automated annotation process led us to identify novel phytochelatin synthase (PCS) genes from two extremophilic green algae (Chlamydomonas acidophila and Dunaliella acidophila). These genes are of intermediate length compared to known PCS genes from eukaryotes and PCS-like genes from prokaryotes. A detailed phylogenetic analysis gives new insight into the complicated evolutionary history of PCS genes and provides evidence for multiple horizontal gene transfer events from bacteria to eukaryotes within the gene family. A separate subgroup containing PCS-like genes within the PCS gene family is not supported since the PCS genes are monophyletic only when the PCS-like genes are included. The presence and functionality of the novel genes in the organisms were verified by genomic sequencing and qRT-PCR. Furthermore, the novel PCS gene in Chlamydomonas acidophila showed very strong induction by cadmium. Cloning and expression of the gene in Escherichia coli clearly improves its cadmium resistance. The gene in Dunaliella was not induced, most likely due to gene duplication.

  7. A pneumatic transfer system for special form {sup 252}Cf

    SciTech Connect

    Gehrke, R.J.; Berry, S.M.; Grafwallner, E.G.; Hoggan, J.M.

    1996-09-01

    A pneumatic transfer system has been developed for use with series 100 Special Form {sup 252}Cf. It was developed to reduce the exposure to personnel handling sources of {sup 252}Cf with masses up to 150 {micro}g by permitting remotely activated two-way transfer between the storage container and the irradiation position. The pneumatic transfer system also permits transfers for reproducible repetitive irradiation periods. In addition to the storage container equipped with quick-release fittings, the transfer system consists of an irradiation station, a control box with momentary contact switches to activate the air-pressure control valves and indicators to identify the location of the source, and connecting air hose and electrical wire. A source of 20 psig air and 110 volt electrical power are required for operation of the transfer system which can be easily moved and set up by one individual in 5 to 10 minutes. Tests have shown that rarely does a source become lodged in the transfer tubing, but two methods have been developed to handle incomplete transfers of the {sup 252}Cf source. The first method consists of closing one air vent to allow a pressure impulse to propel the source to the opposite side. The second method applies to those {sup 252}Cf capsules with a threaded or tapped end to which a small ferromagnetic piece can be attached; an incompletely transferred source in the transfer tube can then be guided to a position of safety by surrounding the transfer tubing containing the capsule with a horseshoe magnet attached to the end of a long pole.

  8. Evidence for Horizontal Gene Transfer as Origin of Putrescine Production in Oenococcus oeni RM83▿

    PubMed Central

    Marcobal, Ángela; de las Rivas, Blanca; Moreno-Arribas, M. Victoria; Muñoz, Rosario

    2006-01-01

    The nucleotide sequence of a 17.2-kb chromosomal DNA fragment containing the odc gene encoding ornithine decarboxylase has been determined in the putrescine producer Oenococcus oeni RM83. This DNA fragment contains 13 open reading frames, including genes coding for five transposases and two phage proteins. This description might represent the first evidence of a horizontal gene transfer event as the origin of a biogenic amine biosynthetic locus. PMID:17056681

  9. Visual prosthesis wireless energy transfer system optimal modeling

    PubMed Central

    2014-01-01

    Background Wireless energy transfer system is an effective way to solve the visual prosthesis energy supply problems, theoretical modeling of the system is the prerequisite to do optimal energy transfer system design. Methods On the basis of the ideal model of the wireless energy transfer system, according to visual prosthesis application condition, the system modeling is optimized. During the optimal modeling, taking planar spiral coils as the coupling devices between energy transmitter and receiver, the effect of the parasitic capacitance of the transfer coil is considered, and especially the concept of biological capacitance is proposed to consider the influence of biological tissue on the energy transfer efficiency, resulting in the optimal modeling’s more accuracy for the actual application. Results The simulation data of the optimal model in this paper is compared with that of the previous ideal model, the results show that under high frequency condition, the parasitic capacitance of inductance and biological capacitance considered in the optimal model could have great impact on the wireless energy transfer system. The further comparison with the experimental data verifies the validity and accuracy of the optimal model proposed in this paper. Conclusions The optimal model proposed in this paper has a higher theoretical guiding significance for the wireless energy transfer system’s further research, and provide a more precise model reference for solving the power supply problem in visual prosthesis clinical application. PMID:24428906

  10. Inhibition of solid tumor growth by gene transfer of VEGF receptor-1 mutants.

    PubMed

    Heidenreich, Regina; Machein, Marcia; Nicolaus, Anke; Hilbig, Andreas; Wild, Carola; Clauss, Matthias; Plate, Karl H; Breier, Georg

    2004-09-01

    Vascular endothelial growth factor (VEGF) and the high-affinity VEGF receptor Flk-1/KDR (VEGFR-2) are key regulators of tumor angiogenesis. Strategies to block VEGF/VEGFR-2 signaling were successfully used to inhibit experimental tumor growth and indicated that VEGFR-2 is the main signaling VEGF receptor in proliferating tumor endothelium. Here, we investigated the role of the VEGF receptor-1 (VEGFR-1/Flt-1) in the vascularization of 2 different experimental tumors in vivo. VEGFR-1 mutants were generated that lack the intracellular tyrosine kinase domain. Retrovirus-mediated gene transfer of the VEGFR-1 mutants led to a strong reduction of tumor growth and angiogenesis in xenografted C6 glioma and in syngeneic BFS-1 fibrosarcoma. Histological analysis of the inhibited fibrosarcoma revealed reduced vascular density, decreased tumor cell proliferation as well as increased tumor cell apoptosis and the formation of necrosis. The retroviral gene transfer of the full length VEGFR-1 also caused a significant reduction of tumor growth in both models. The inhibitory effects of the VEGFR-1 mutants and the full length VEGFR-1 in BFS-1 fibrosarcoma were mediated through host tumor endothelial cells because the BFS-1 fibrosarcoma cells were not infected by the retrovirus. The formation of heterodimers between VEGFR-2 and full length or truncated VEGFR-1 was observed in vitro and might contribute to the growth inhibitory effect by modulating distinct signal transduction pathways. The results of our study underline the central role of the VEGF/VEGFR-1 signaling system in tumor angiogenesis and demonstrate that VEGFR-1 can serve as a target for anti-angiogenic gene therapy.

  11. No evidence of inhibition of horizontal gene transfer by CRISPR-Cas on evolutionary timescales.

    PubMed

    Gophna, Uri; Kristensen, David M; Wolf, Yuri I; Popa, Ovidiu; Drevet, Christine; Koonin, Eugene V

    2015-09-01

    The CRISPR (clustered, regularly, interspaced, short, palindromic repeats)-Cas (CRISPR-associated genes) systems of archaea and bacteria provide adaptive immunity against viruses and other selfish elements and are believed to curtail horizontal gene transfer (HGT). Limiting acquisition of new genetic material could be one of the sources of the fitness cost of CRISPR-Cas maintenance and one of the causes of the patchy distribution of CRISPR-Cas among bacteria, and across environments. We sought to test the hypothesis that the activity of CRISPR-Cas in microbes is negatively correlated with the extent of recent HGT. Using three independent measures of HGT, we found no significant dependence between the length of CRISPR arrays, which reflects the activity of the immune system, and the estimated number of recent HGT events. In contrast, we observed a significant negative dependence between the estimated extent of HGT and growth temperature of microbes, which could be explained by the lower genetic diversity in hotter environments. We hypothesize that the relevant events in the evolution of resistance to mobile elements and proclivity for HGT, to which CRISPR-Cas systems seem to substantially contribute, occur on the population scale rather than on the timescale of species evolution.

  12. Gene Loss and Horizontal Gene Transfer Contributed to the Genome Evolution of the Extreme Acidophile "Ferrovum".

    PubMed

    Ullrich, Sophie R; González, Carolina; Poehlein, Anja; Tischler, Judith S; Daniel, Rolf; Schlömann, Michael; Holmes, David S; Mühling, Martin

    2016-01-01

    Acid mine drainage (AMD), associated with active and abandoned mining sites, is a habitat for acidophilic microorganisms that gain energy from the oxidation of reduced sulfur compounds and ferrous iron and that thrive at pH below 4. Members of the recently proposed genus "Ferrovum" are the first acidophilic iron oxidizers to be described within the Betaproteobacteria. Although they have been detected as typical community members in AMD habitats worldwide, knowledge of their phylogenetic and metabolic diversity is scarce. Genomics approaches appear to be most promising in addressing this lacuna since isolation and cultivation of "Ferrovum" has proven to be extremely difficult and has so far only been successful for the designated type strain "Ferrovum myxofaciens" P3G. In this study, the genomes of two novel strains of "Ferrovum" (PN-J185 and Z-31) derived from water samples of a mine water treatment plant were sequenced. These genomes were compared with those of "Ferrovum" sp. JA12 that also originated from the mine water treatment plant, and of the type strain (P3G). Phylogenomic scrutiny suggests that the four strains represent three "Ferrovum" species that cluster in two groups (1 and 2). Comprehensive analysis of their predicted metabolic pathways revealed that these groups harbor characteristic metabolic profiles, notably with respect to motility, chemotaxis, nitrogen metabolism, biofilm formation and their potential strategies to cope with the acidic environment. For example, while the "F. myxofaciens" strains (group 1) appear to be motile and diazotrophic, the non-motile group 2 strains have the predicted potential to use a greater variety of fixed nitrogen sources. Furthermore, analysis of their genome synteny provides first insights into their genome evolution, suggesting that horizontal gene transfer and genome reduction in the group 2 strains by loss of genes encoding complete metabolic pathways or physiological features contributed to the observed

  13. Horizontal Transfer and Death of a Fungal Secondary Metabolic Gene Cluster

    PubMed Central

    Campbell, Matthew A.; Rokas, Antonis; Slot, Jason C.

    2012-01-01

    A cluster composed of four structural and two regulatory genes found in several species of the fungal genus Fusarium (class Sordariomycetes) is responsible for the production of the red pigment bikaverin. We discovered that the unrelated fungus Botrytis cinerea (class Leotiomycetes) contains a cluster of five genes that is highly similar in sequence and gene order to the Fusarium bikaverin cluster. Synteny conservation, nucleotide composition, and phylogenetic analyses of the cluster genes indicate that the B. cinerea cluster was acquired via horizontal transfer from a Fusarium donor. Upon or subsequent to the transfer, the B. cinerea gene cluster became inactivated; one of the four structural genes is missing, two others are pseudogenes, and the fourth structural gene shows an accelerated rate of nonsynonymous substitutions along the B. cinerea lineage, consistent with relaxation of selective constraints. Interestingly, the bik4 regulatory gene is still intact and presumably functional, whereas bik5, which is a pathway-specific regulator, also shows a mild but significant acceleration of evolutionary rate along the B. cinerea lineage. This selective preservation of the bik4 regulator suggests that its conservation is due to its likely involvement in other non–bikaverin-related biological processes in B. cinerea. Thus, in addition to novel metabolism, horizontal transfer of wholesale metabolic gene clusters might also be contributing novel regulation. PMID:22294497

  14. Optimizing A Lipocomplex-Based Gene Transfer Method into HeLa Cell Line

    PubMed Central

    Asgharian, Alimohammad; Banan, Mehdi; Najmabadi, Hossein

    2014-01-01

    One of the most significant steps in gene expression studies is transferring genes into cell cultures. Despite there are different methods for gene delivery such as viral and non-viral producers, some cationic lipid reagents have recently developed to transfect into mam- malian cell lines. The main aim of this study was optimizing and improving lipocomplex based transient transfection procedures into HeLa cell line which is being used widely as a typical cell in biological studies. This study was an experimental research. In this work, pCMV β-Gal DNA plasmid was used as a reporter DNA for determining the rate of gene transfection into HeLa cells. To accomplish the highest gene delivery into HeLa cells, optimizing experiments were carried out in different volumes of FuGENE-HD, LipofectamineTM2000 and X-tremeGENE. Also, we investigated tranasfection efficiency in presence of various cell densities of HeLa cells. Then, transfection efficiency and cell toxicity were measured by beta gal staining and trypan blue methods, respectively. Using FuGENE-HD in volume of 4µl along with 105 HeLa cells, transfection efficiency was higher (43.66 ± 1.52%) in comparison with the cationic lipids LipofectamineTM2000 and X-tremeGENE. In addition, the rate of cell toxicity in presence of FuGENE-HD was less than 5%. In summary, the cationic lipid FuGENE-HD indicates a suitable potential to transfer DNA into HeLa cells and it can be an efficient reagent for gene delivery for HeLa cells in vitro. Moreover, it is worth designing and optimizing gene transfer experiments for other cell lines with FuGENE-HD due to its low toxicity and high efficiency. PMID:24381863

  15. Optimizing A Lipocomplex-Based Gene Transfer Method into HeLa Cell Line.

    PubMed

    Asgharian, Alimohammad; Banan, Mehdi; Najmabadi, Hossein

    2014-01-01

    One of the most significant steps in gene expression studies is transferring genes into cell cultures. Despite there are different methods for gene delivery such as viral and non-viral producers, some cationic lipid reagents have recently developed to transfect into mam- malian cell lines. The main aim of this study was optimizing and improving lipocomplex based transient transfection procedures into HeLa cell line which is being used widely as a typical cell in biological studies. This study was an experimental research. In this work, pCMV β-Gal DNA plasmid was used as a reporter DNA for determining the rate of gene transfection into HeLa cells. To accomplish the highest gene delivery into HeLa cells, optimizing experiments were car- ried out in different volumes of FuGENE-HD, Lipofectamine(TM)2000 and X-tremeGENE. Also, we investigated tranasfection efficiency in presence of various cell densities of HeLa cells. Then, transfection efficiency and cell toxicity were measured by beta gal staining and trypan blue methods, respectively. Using FuGENE-HD in volume of 4µl along with 10(5) HeLa cells, transfection efficiency was higher (43.66 ± 1.52%) in comparison with the cationic lipids Lipofectamine(TM)2000 and X-tremeGENE. In addition, the rate of cell toxicity in presence of FuGENE-HD was less than 5%. In summary, the cationic lipid FuGENE-HD indicates a suitable potential to transfer DNA into HeLa cells and it can be an efficient reagent for gene delivery for HeLa cells in vitro. Moreover, it is worth designing and optimizing gene transfer experiments for other cell lines with FuGENE-HD due to its low toxicity and high efficiency.

  16. Guanidinylated block copolymers for gene transfer: A comparison with amine-based materials for in vitro and in vivo gene transfer efficiency

    PubMed Central

    Choi, Jennifer L.; Tan, James-Kevin Y.; Sellers, Drew L.; Wei, Hua; Horner, Philip J.; Pun, Suzie H.

    2015-01-01

    There is currently no cure for neuron loss in the brain, which can occur due to traumatic injury or neurodegenerative disease. One method proposed to enhance neurogenesis in the brain is gene transfer to neural progenitor cells. In this work, a guanidine-based copolymer was synthesized and compared to an amine-based copolymer analog previously shown to effectively deliver genes in the murine brain. The guanidine-based copolymer was more efficient at gene transfer to immortalized, cultured cell lines; however, the amine-based copolymer was more effective at gene transfer in the brain. DNA condensation studies revealed that the nucleic acid complexes formed with the guanidine-based copolymer were more susceptible to unpackaging in the presence of heparin sulfate proteoglycans compared to complexes formed with the amine-based copolymer. Therefore, polyplexes formed from the amine-based copolymer may be more resistant to destabilization by the heparan sulfate proteoglycans present in the stem cell niches of the brain. PMID:25907042

  17. Two Horizontally Transferred Xenobiotic Resistance Gene Clusters Associated with Detoxification of Benzoxazolinones by Fusarium Species

    PubMed Central

    Glenn, Anthony E.; Davis, C. Britton; Gao, Minglu; Gold, Scott E.; Mitchell, Trevor R.; Proctor, Robert H.; Stewart, Jane E.; Snook, Maurice E.

    2016-01-01

    Microbes encounter a broad spectrum of antimicrobial compounds in their environments and often possess metabolic strategies to detoxify such xenobiotics. We have previously shown that Fusarium verticillioides, a fungal pathogen of maize known for its production of fumonisin mycotoxins, possesses two unlinked loci, FDB1 and FDB2, necessary for detoxification of antimicrobial compounds produced by maize, including the γ-lactam 2-benzoxazolinone (BOA). In support of these earlier studies, microarray analysis of F. verticillioides exposed to BOA identified the induction of multiple genes at FDB1 and FDB2, indicating the loci consist of gene clusters. One of the FDB1 cluster genes encoded a protein having domain homology to the metallo-β-lactamase (MBL) superfamily. Deletion of this gene (MBL1) rendered F. verticillioides incapable of metabolizing BOA and thus unable to grow on BOA-amended media. Deletion of other FDB1 cluster genes, in particular AMD1 and DLH1, did not affect BOA degradation. Phylogenetic analyses and topology testing of the FDB1 and FDB2 cluster genes suggested two horizontal transfer events among fungi, one being transfer of FDB1 from Fusarium to Colletotrichum, and the second being transfer of the FDB2 cluster from Fusarium to Aspergillus. Together, the results suggest that plant-derived xenobiotics have exerted evolutionary pressure on these fungi, leading to horizontal transfer of genes that enhance fitness or virulence. PMID:26808652

  18. Two Horizontally Transferred Xenobiotic Resistance Gene Clusters Associated with Detoxification of Benzoxazolinones by Fusarium Species.

    PubMed

    Glenn, Anthony E; Davis, C Britton; Gao, Minglu; Gold, Scott E; Mitchell, Trevor R; Proctor, Robert H; Stewart, Jane E; Snook, Maurice E

    2016-01-01

    Microbes encounter a broad spectrum of antimicrobial compounds in their environments and often possess metabolic strategies to detoxify such xenobiotics. We have previously shown that Fusarium verticillioides, a fungal pathogen of maize known for its production of fumonisin mycotoxins, possesses two unlinked loci, FDB1 and FDB2, necessary for detoxification of antimicrobial compounds produced by maize, including the γ-lactam 2-benzoxazolinone (BOA). In support of these earlier studies, microarray analysis of F. verticillioides exposed to BOA identified the induction of multiple genes at FDB1 and FDB2, indicating the loci consist of gene clusters. One of the FDB1 cluster genes encoded a protein having domain homology to the metallo-β-lactamase (MBL) superfamily. Deletion of this gene (MBL1) rendered F. verticillioides incapable of metabolizing BOA and thus unable to grow on BOA-amended media. Deletion of other FDB1 cluster genes, in particular AMD1 and DLH1, did not affect BOA degradation. Phylogenetic analyses and topology testing of the FDB1 and FDB2 cluster genes suggested two horizontal transfer events among fungi, one being transfer of FDB1 from Fusarium to Colletotrichum, and the second being transfer of the FDB2 cluster from Fusarium to Aspergillus. Together, the results suggest that plant-derived xenobiotics have exerted evolutionary pressure on these fungi, leading to horizontal transfer of genes that enhance fitness or virulence.

  19. Carotenoids in unexpected places: gall midges, lateral gene transfer, and carotenoid biosynthesis in animals.

    PubMed

    Cobbs, Cassidy; Heath, Jeremy; Stireman, John O; Abbot, Patrick

    2013-08-01

    Carotenoids are conjugated isoprenoid molecules with many important physiological functions in organisms, including roles in photosynthesis, oxidative stress reduction, vision, diapause, photoperiodism, and immunity. Until recently, it was believed that only plants, microorganisms, and fungi were capable of synthesizing carotenoids and that animals acquired them from their diet, but recent studies have demonstrated that two arthropods (pea aphid and spider mite) possess a pair of genes homologous to those required for the first step of carotenoid biosynthesis. Absent in all other known animal genomes, these genes appear to have been acquired by aphids and spider mites in one or several lateral gene transfer events from a fungal donor. We report the third case of fungal carotenoid biosynthesis gene homologs in an arthropod: flies from the family Cecidomyiidae, commonly known as gall midges. Using phylogenetic analyses we show that it is unlikely that lycopene cyclase/phytoene synthase and phytoene desaturase homologs were transferred singly to an ancient arthropod ancestor; instead we propose that genes were transferred independently from related fungal donors after divergence of the major arthropod lineages. We also examine variation in intron placement and copy number of the carotenoid genes that may underlie function in the midges. This trans-kingdom transfer of carotenoid genes may represent a key innovation, underlying the evolution of phytophagy and plant-galling in gall midges and facilitating their extensive diversification across plant lineages.

  20. CD133-targeted gene transfer into long-term repopulating hematopoietic stem cells.

    PubMed

    Brendel, Christian; Goebel, Benjamin; Daniela, Abriss; Brugman, Martijn; Kneissl, Sabrina; Schwäble, Joachim; Kaufmann, Kerstin B; Müller-Kuller, Uta; Kunkel, Hana; Chen-Wichmann, Linping; Abel, Tobias; Serve, Hubert; Bystrykh, Leonid; Buchholz, Christian J; Grez, Manuel

    2015-01-01

    Gene therapy for hematological disorders relies on the genetic modification of CD34(+) cells, a heterogeneous cell population containing about 0.01% long-term repopulating cells. Here, we show that the lentiviral vector CD133-LV, which uses a surface marker on human primitive hematopoietic stem cells (HSCs) as entry receptor, transfers genes preferentially into cells with high engraftment capability. Transduction of unstimulated CD34(+) cells with CD133-LV resulted in gene marking of cells with competitive proliferative advantage in vitro and in immunodeficient mice. The CD133-LV-transduced population contained significantly more cells with repopulating capacity than cells transduced with vesicular stomatitis virus (VSV)-LV, a lentiviral vector pseudotyped with the vesicular stomatitis virus G protein. Upon transfer of a barcode library, CD133-LV-transduced cells sustained gene marking in vivo for a prolonged period of time with a 6.7-fold higher recovery of barcodes compared to transduced control cells. Moreover, CD133-LV-transduced cells were capable of repopulating secondary recipients. Lastly, we show that this targeting strategy can be used for transfer of a therapeutic gene into CD34(+) cells obtained from patients suffering of X-linked chronic granulomatous disease. In conclusion, direct gene transfer into CD133(+) cells allows for sustained long-term engraftment of gene corrected cells.

  1. Mycelia as a focal point for horizontal gene transfer among soil bacteria

    PubMed Central

    Berthold, Tom; Centler, Florian; Hübschmann, Thomas; Remer, Rita; Thullner, Martin; Harms, Hauke; Wick, Lukas Y.

    2016-01-01

    Horizontal gene transfer (HGT) is a main mechanism of bacterial evolution endowing bacteria with new genetic traits. The transfer of mobile genetic elements such as plasmids (conjugation) requires the close proximity of cells. HGT between genetically distinct bacteria largely depends on cell movement in water films, which are typically discontinuous in natural systems like soil. Using laboratory microcosms, a bacterial reporter system and flow cytometry, we here investigated if and to which degree mycelial networks facilitate contact of and HGT between spatially separated bacteria. Our study shows that the network structures of mycelia promote bacterial HGT by providing continuous liquid films in which bacterial migration and contacts are favoured. This finding was confirmed by individual-based simulations, revealing that the tendency of migrating bacteria to concentrate in the liquid film around hyphae is a key factor for improved HGT along mycelial networks. Given their ubiquity, we propose that hyphae can act as focal point for HGT and genetic adaptation in soil. PMID:27811990

  2. Phonon-assisted excitation energy transfer in photosynthetic systems

    NASA Astrophysics Data System (ADS)

    Chen, Hao; Wang, Xin; Fang, Ai-Ping; Li, Hong-Rong

    2016-09-01

    The phonon-assisted process of energy transfer aiming at exploring the newly emerging frontier between biology and physics is an issue of central interest. This article shows the important role of the intramolecular vibrational modes for excitation energy transfer in the photosynthetic systems. Based on a dimer system consisting of a donor and an acceptor modeled by two two-level systems, in which one of them is coupled to a high-energy vibrational mode, we derive an effective Hamiltonian describing the vibration-assisted coherent energy transfer process in the polaron frame. The effective Hamiltonian reveals in the case that the vibrational mode dynamically matches the energy detuning between the donor and the acceptor, the original detuned energy transfer becomes resonant energy transfer. In addition, the population dynamics and coherence dynamics of the dimer system with and without vibration-assistance are investigated numerically. It is found that, the energy transfer efficiency and the transfer time depend heavily on the interaction strength of the donor and the high-energy vibrational mode, as well as the vibrational frequency. The numerical results also indicate that the initial state and dissipation rate of the vibrational mode have little influence on the dynamics of the dimer system. Results obtained in this article are not only helpful to understand the natural photosynthesis, but also offer an optimal design principle for artificial photosynthesis. Project supported by the National Natural Science Foundation of China (Grant No. 11174233).

  3. In vitro assembly of a prohead-like structure of the Rhodobacter capsulatus gene transfer agent

    SciTech Connect

    Spano, Anthony J. . E-mail: ajs6z@virginia.edu; Chen, Frank S.; Goodman, Benjamin E.; Sabat, Agnes E.; Simon, Martha N.; Wall, Joseph S.; Correia, John J.; McIvor, Wilson; Newcomb, William W.; Brown, Jay C.; Schnur, Joel M.; Lebedev, Nikolai

    2007-07-20

    The gene transfer agent (GTA) is a phage-like particle capable of exchanging double-stranded DNA fragments between cells of the photosynthetic bacterium Rhodobacter capsulatus. Here we show that the major capsid protein of GTA, expressed in E. coli, can be assembled into prohead-like structures in the presence of calcium ions in vitro. Transmission electron microscopy (TEM) of uranyl acetate staining material and thin sections of glutaraldehyde-fixed material demonstrates that these associates have spherical structures with diameters in the range of 27-35 nm. The analysis of scanning TEM images revealed particles of mass {approx} 4.3 MDa, representing 101 {+-} 11 copies of the monomeric subunit. The establishment of this simple and rapid method to form prohead-like particles permits the GTA system to be used for genome manipulation within the photosynthetic bacterium, for specific targeted drug delivery, and for the construction of biologically based distributed autonomous sensors for environmental monitoring.

  4. Horizontal transfer of archaeal genes into the deinococcaceae: detection by molecular and computer-based approaches

    NASA Technical Reports Server (NTRS)

    Olendzenski, L.; Liu, L.; Zhaxybayeva, O.; Murphey, R.; Shin, D. G.; Gogarten, J. P.

    2000-01-01

    Members of the Deinococcaceae (e.g., Thermus, Meiothermus, Deinococcus) contain A/V-ATPases typically found in Archaea or Eukaryotes which were probably acquired by horizontal gene transfer. Two methods were used to quantify the extent to which archaeal or eukaryotic genes have been acquired by this lineage. Screening of a Meiothermus ruber library with probes made against Thermoplasma acidophilum DNA yielded a number of clones which hybridized more strongly than background. One of these contained the prolyl tRNA synthetase (RS) gene. Phylogenetic analysis shows the M. ruber and D. radiodurans prolyl RS to be more closely related to archaeal and eukaryal forms of this gene than to the typical bacterial type. Using a bioinformatics approach, putative open reading frames (ORFs) from the prerelease version of the D. radiodurans genome were screened for genes more closely related to archaeal or eukaryotic genes. Putative ORFs were searched against representative genomes from each of the three domains using automated BLAST. ORFs showing the highest matches against archaeal and eukaryotic genes were collected and ranked. Among the top-ranked hits were the A/V-ATPase catalytic and noncatalytic subunits and the prolyl RS genes. Using phylogenetic methods, ORFs were analyzed and trees assessed for evidence of horizontal gene transfer. Of the 45 genes examined, 20 showed topologies in which D. radiodurans homologues clearly group with eukaryotic or archaeal homologues, and 17 additional trees were found to show probable evidence of horizontal gene transfer. Compared to the total number of ORFs in the genome, those that can be identified as having been acquired from Archaea or Eukaryotes are relatively few (approximately 1%), suggesting that interdomain transfer is rare.

  5. Generation of hypoxanthine phosphoribosyltransferase gene knockout rabbits by homologous recombination and gene trapping through somatic cell nuclear transfer.

    PubMed

    Yin, Mingru; Jiang, Weihua; Fang, Zhenfu; Kong, Pengcheng; Xing, Fengying; Li, Yao; Chen, Xuejin; Li, Shangang

    2015-11-02

    The rabbit is a common animal model that has been employed in studies on various human disorders, and the generation of genetically modified rabbit lines is highly desirable. Female rabbits have been successfully cloned from cumulus cells, and the somatic cell nuclear transfer (SCNT) technology is well established. The present study generated hypoxanthine phosphoribosyltransferase (HPRT) gene knockout rabbits using recombinant adeno-associated virus-mediated homologous recombination and SCNT. Gene trap strategies were employed to enhance the gene targeting rates. The male and female gene knockout fibroblast cell lines were derived by different strategies. When male HPRT knockout cells were used for SCNT, no live rabbits were obtained. However, when female HPRT(+/-) cells were used for SCNT, live, healthy rabbits were generated. The cloned HPRT(+/-) rabbits were fertile at maturity. We demonstrate a new technique to produce gene-targeted rabbits. This approach may also be used in the genetic manipulation of different genes or in other species.

  6. Criticality Evaluation - Cask Unloading Using the Shielded Transfer System

    SciTech Connect

    Blanchard, A.; Nadeau, M.L.

    1998-06-01

    This evaluation reviewed the criticality aspects of Shielded Transfer System (STS) operation. The existing Nuclear Safety Control (NSC) elements were examined and new NSC elements were developed, as needed.

  7. 40 CFR 63.689 - Standards: Transfer systems.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... in the transfer system; the effects of outdoor exposure to wind, moisture, and sunlight; and the... suitable materials that will minimize exposure of the off-site material to the atmosphere, to the...

  8. High Quality Factor Resonators for Inductive Power Transfer Systems

    NASA Astrophysics Data System (ADS)

    Etemadrezaei, Mohammad

    In this dissertation, the Inductive Power Transfer (IPT) systems for multi-MHz frequency of operation are investigated, and new ideas for magnetic link inductive coils are presented. (Abstract shortened by ProQuest.).

  9. Investigation of horizontal gene transfer in poplar/Amanita muscaria ectomycorrhizas.

    PubMed

    Zhang, Chi; Hampp, Rüdiger; Nehls, Uwe

    2005-01-01

    Fine roots of forest trees form together with certain soil fungi symbiotic structures (ectomycorrhizas), where fungal hyphae are in intimate contact with plant cells. Due to root cell degeneration, plant DNA is released and could be taken up by the fungus. The possibility that horizontal gene transfer might result in a risk for the environment should be evaluated before a massive release of genetically engineered trees into nature occurs, even though only a few convincing examples of horizontal gene transfer are known. Transgenic poplars containing a construct of the Streptomyces hygroscopicus bar gene under the control of the Cochliobolus heterostrophus GPD (glyceraldehyde-3-phosphate dehydrogenase) promoter were generated by Agrobacterium-mediated transformation. The functionality of this construct in the ectomycorrhizal model fungus Amanita muscaria was previously verified by protoplast-based fungal transformation. 35,000 ectomycorrhizas, formed between transgenic poplars and non-transgenic A. muscaria hyphae, were isolated and transferred to selective agar plates. Putative herbicide-resistant fungal colonies were obtained after the first round of selection. However, none of these colonies survived a transfer onto fresh selection medium, nor did they contain the bar gene, indicating that no horizontal gene transfer from poplar to A. muscaria occurred during symbiosis under axenic conditions. However, since ectomycorrhizas are associated under natural conditions with viruses, bacteria and other fungi, these additional associations should be evaluated in future.

  10. Plant expansins in bacteria and fungi: evolution by horizontal gene transfer and independent domain fusion.

    PubMed

    Nikolaidis, Nikolas; Doran, Nicole; Cosgrove, Daniel J

    2014-02-01

    Horizontal gene transfer (HGT) has been described as a common mechanism of transferring genetic material between prokaryotes, whereas genetic transfers from eukaryotes to prokaryotes have been rarely documented. Here we report a rare case of HGT in which plant expansin genes that code for plant cell-wall loosening proteins were transferred from plants to bacteria, fungi, and amoebozoa. In several cases, the species in which the expansin gene was found is either in intimate association with plants or is a known plant pathogen. Our analyses suggest that at least two independent genetic transfers occurred from plants to bacteria and fungi. These events were followed by multiple HGT events within bacteria and fungi. We have also observed that in bacteria expansin genes have been independently fused to DNA fragments that code for an endoglucanase domain or for a carbohydrate binding module, pointing to functional convergence at the molecular level. Furthermore, the functional similarities between microbial expansins and their plant xenologs suggest that these proteins mediate microbial-plant interactions by altering the plant cell wall and therefore may provide adaptive advantages to these species. The evolution of these nonplant expansins represents a unique case in which bacteria and fungi have found innovative and adaptive ways to interact with and infect plants by acquiring genes from their host. This evolutionary paradigm suggests that despite their low frequency such HGT events may have significantly contributed to the evolution of prokaryotic and eukaryotic species.

  11. AAV8-mediated hepatic gene transfer in infant rhesus monkeys (Macaca mulatta).

    PubMed

    Wang, Lili; Bell, Peter; Lin, Jianping; Calcedo, Roberto; Tarantal, Alice F; Wilson, James M

    2011-11-01

    Many genetic metabolic diseases manifest in infancy, therefore, it is important to develop effective treatments that could be implemented at this time. Adeno-associated virus serotype 8 (AAV8) gene transfer has been studied in neonatal mouse, cat, and dog models and shown some efficacy with a single hepatic injection at birth. AAV8-mediated liver gene transfer has also generated sustained therapeutic effects in feline and canine models of lysosomal storage disorders. In these models, delaying the age of vector treatment increased gene transfer stability. The growth rate of infant nonhuman primates is more similar to the growth trajectory of humans, thus infant monkeys provide an excellent model to study AAV gene transfer efficiency, stability, and safety. In this study, we report for the first time that AAV8-mediated hepatic gene transfer in infant monkeys is safe and efficient but less stable when compared to adolescent animals. Infant monkeys administered AAV8 intravenously at 1 week postnatal age achieved up to 98% transduction of hepatocytes within 7 days of injection; however, there was significant dilution of genomes and loss of transgene expression 35 days postadministration. Delaying the injection to 1 month postnatal age did not improve stability of transduction but decreased the antibody response to AAV8 capsid.

  12. Transferring Sclerotinia Resistance Genes from Wild Helianthus into Cultivated Sunflower

    Technology Transfer Automated Retrieval System (TEKTRAN)

    To enhance resistance to Sclerotinia head and stalk rot in cultivated sunflower, mining and introgression of Sclerotinia resistance genes from diverse wild Helianthus accessions into cultivated sunflower has been conducted using backcrossing method since 2004. During the last four years, numerous in...

  13. Efficient methods for finding transfer function zeros of power systems

    SciTech Connect

    Martins, N.; Pinto, H.J.C.P. ); Lima, L.T.G. )

    1992-08-01

    This paper is probably the first to describe algorithms suited to the efficient calculation of both proper and non-proper transfer function zeros of linearized dynamic models for large interconnected power systems. The paper also describes an improvement to the well known AESOPS algorithm, formulating it as an exact transfer function zero finding problem which is efficiently solved by a Newton-Raphson iterative scheme. Large power system results are presented in this paper.

  14. Chemoreceptor Gene Loss and Acquisition via Horizontal Gene Transfer in Escherichia coli

    PubMed Central

    Borziak, Kirill; Fleetwood, Aaron D.

    2013-01-01

    Chemotaxis allows bacteria to more efficiently colonize optimal microhabitats within their larger environment. Chemotaxis in Escherichia coli is the best-studied model system, and a large number of E. coli strains have been sequenced. The Escherichia/Shigella genus encompasses a great variety of commensal and pathogenic strains, but the role of chemotaxis in their association with the host remains poorly understood. Here we show that the core chemotaxis genes are lost in many, but not all, nonmotile strains but are well preserved in all motile strains. The genes encoding the Tar, Tsr, and Aer chemoreceptors, which mediate chemotaxis to a broad spectrum of chemical and physical cues, are also nearly uniformly conserved in motile strains. In contrast, the clade of extraintestinal pathogenic E. coli strains apparently underwent an ancestral loss of Trg and Tap chemoreceptors, which sense sugars, dipeptides, and pyrimidines. The broad range of time estimated for the loss of these genes (1 to 3 million years ago) corresponds to the appearance of the genus Homo. PMID:23749975

  15. Massive gene transfer and extensive RNA editing of a symbiotic dinoflagellate plastid genome.

    PubMed

    Mungpakdee, Sutada; Shinzato, Chuya; Takeuchi, Takeshi; Kawashima, Takeshi; Koyanagi, Ryo; Hisata, Kanako; Tanaka, Makiko; Goto, Hiroki; Fujie, Manabu; Lin, Senjie; Satoh, Nori; Shoguchi, Eiichi

    2014-05-31

    Genome sequencing of Symbiodinium minutum revealed that 95 of 109 plastid-associated genes have been transferred to the nuclear genome and subsequently expanded by gene duplication. Only 14 genes remain in plastids and occur as DNA minicircles. Each minicircle (1.8-3.3 kb) contains one gene and a conserved noncoding region containing putative promoters and RNA-binding sites. Nine types of RNA editing, including a novel G/U type, were discovered in minicircle transcripts but not in genes transferred to the nucleus. In contrast to DNA editing sites in dinoflagellate mitochondria, which tend to be highly conserved across all taxa, editing sites employed in DNA minicircles are highly variable from species to species. Editing is crucial for core photosystem protein function. It restores evolutionarily conserved amino acids and increases peptidyl hydropathy. It also increases protein plasticity necessary to initiate photosystem complex assembly.

  16. Mucus altering agents as adjuncts for nonviral gene transfer to airway epithelium.

    PubMed

    Ferrari, S; Kitson, C; Farley, R; Steel, R; Marriott, C; Parkins, D A; Scarpa, M; Wainwright, B; Evans, M J; Colledge, W H; Geddes, D M; Alton, E W

    2001-09-01

    Nonviral vectors have been shown to be a safe and valid alternative to recombinant viruses for gene therapy of cystic fibrosis (CF). Nevertheless, gene transfer efficiency needs to be increased before clinical efficacy is likely in man. One barrier to increased efficacy is normal airway mucus. Using an ex vivo model of sheep tracheal epithelium, we show that this barrier can, in part, be overcome by treatment with the mucolytic agents, Nacystelyn or N-acetylcysteine using either a cationic lipid or a cationic polymer as the gene transfer agent. Further, in vivo application of either Nacystelyn or the anticholinergic glycopyrrolate, both clinically used agents, resulted in increased reporter gene expression in the mouse lung, but no significant correction of the bioelectric defect in CF null mice. These results, whilst unlikely to be sufficient in themselves to achieve clinically relevant gene therapy, may be a further useful step in the attainment of this goal.

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

    SciTech Connect

    Marton, L.

    1994-12-31

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

  18. Lines of evidence for horizontal gene transfer of a phenazine producing operon into multiple bacterial species.

    PubMed

    Fitzpatrick, David A

    2009-02-01

    Phenazines are secondary metabolites with broad-spectrum antibiotic activity against bacteria, fungi, and eukaryotes. In pseudomonad species, a conserved seven-gene phenazine operon (phzABCDEFG) is required for the conversion of chorismic acid to the broad-spectrum antibiotic phenazine-1-carboxylate. Previous analyses of genes involved in phenazine production from nonpseudomonad species uncovered a high degree of sequence similarity to pseudomonad homologues. The analyses undertaken in this study wished to eluciadate the evolutionary history of genes involved in the production of phenazines. Furthermore, I wanted to determine if the phenazine operon has been transferred through horizontal gene transfer. Analyses of GC content, codon usage patterns, frequency of 3:1 dinucleotides, sequence similarities, and phylogenetic reconstructions were undertaken to map the evolutionary history of phenazine genes from multiple bacterial species. Patchy phyletic distribution, high sequence similarities, and phylogenetic evidence infer that pseudomonad, Streptomyces cinnamonensis, Pantoea agglomerans, Burkholderia cepacia, Pectobacterium atrosepticum, Brevibacterium linens, and Mycobacterium abscessus species all contain a phenazine operon which has most likely been transferred among these species through horizontal gene transfer. The acquisition of an antibiotic-associated operon is significant, as it may increase the relative fitness of the recipient species.

  19. Design of auto-tuning capacitive power transfer system for wireless power transfer

    NASA Astrophysics Data System (ADS)

    Lu, Kai; Kiong Nguang, Sing

    2016-09-01

    This paper presents the design of capacitive wireless power transfer systems based on a Class-E inverter approach. The main reason for adopting the Class-E inverter approach is because of its high efficiency, theoretically 100%. However, the operation of a Class-E inverter is highly sensitive to its circuit's parameters. In a typical capacitive wireless power transfer application, the capacitive coupling distance between plates is subject to changes, and hence its power transfer efficiency is greatly affected if the Class-E inverter is properly tuned. This drawback motivates us to develop an auto frequency tuning algorithm for a Class-E inverter which maintains its power transfer efficiency in spite of the variations of capacitive coupling distances between plates and circuit's parameters. Finally, simulation and experiment are carried out to verify the effectiveness of the auto frequency tuning algorithm.

  20. Direct phylogenetic evidence for lateral transfer of elongation factor-like gene.

    PubMed

    Kamikawa, Ryoma; Inagaki, Yuji; Sako, Yoshihiko

    2008-05-13

    Genes encoding elongation factor-like (EFL) proteins, which show high similarity to elongation factor-1alpha (EF-1alpha), have been found in phylogenetically distantly related eukaryotes. The sporadic distribution of "EFL-containing" lineages within "EF-1alpha-containing" lineages indirectly, but strongly, suggests lateral gene transfer as the principal driving force in EFL evolution. However, one of the most critical aspects in the above hypothesis, the donor lineages in any putative cases of lateral EFL gene transfer, remained unclear. In this study, we provide direct evidence for lateral transfer of an EFL gene through the analyses of 10 diatom EFL genes. All diatom EFL homologues tightly clustered in phylogenetic analyses, suggesting acquisition of the exogenous EFL gene early in diatom evolution. Our survey additionally identified Thalassiosira pseudonana as a eukaryote bearing EF-1alpha and EFL genes and secondary EFL gene loss in Phaeodactylum tricornutum, the complete genome of which encodes only the EF-1alpha gene. Most importantly, the EFL phylogeny recovered a robust grouping of homologues from diatoms, the cercozoan Bigelowiella natans, and the foraminifer Planoglabratella opecularis, with the diatoms nested within the Bigelowiella plus Planoglabratella (Rhizaria) grouping. The particular relationships recovered are further consistent with two characteristic sequence motifs. The best explanation of our data analyses is an EFL gene transfer from a foraminifer to a diatom, the first case in which the donor-recipient relationship was clarified. Finally, based on a reverse transcriptase quantitative PCR assay and the genome information of Thalassiosira and Phaeodactylum, we propose the loss of elongation factor function in Thalassiosira EF-1alpha.

  1. Direct phylogenetic evidence for lateral transfer of elongation factor-like gene

    PubMed Central

    Kamikawa, Ryoma; Inagaki, Yuji; Sako, Yoshihiko

    2008-01-01

    Genes encoding elongation factor-like (EFL) proteins, which show high similarity to elongation factor-1α (EF-1α), have been found in phylogenetically distantly related eukaryotes. The sporadic distribution of “EFL-containing” lineages within “EF-1α-containing” lineages indirectly, but strongly, suggests lateral gene transfer as the principal driving force in EFL evolution. However, one of the most critical aspects in the above hypothesis, the donor lineages in any putative cases of lateral EFL gene transfer, remained unclear. In this study, we provide direct evidence for lateral transfer of an EFL gene through the analyses of 10 diatom EFL genes. All diatom EFL homologues tightly clustered in phylogenetic analyses, suggesting acquisition of the exogenous EFL gene early in diatom evolution. Our survey additionally identified Thalassiosira pseudonana as a eukaryote bearing EF-1α and EFL genes and secondary EFL gene loss in Phaeodactylum tricornutum, the complete genome of which encodes only the EF-1α gene. Most importantly, the EFL phylogeny recovered a robust grouping of homologues from diatoms, the cercozoan Bigelowiella natans, and the foraminifer Planoglabratella opecularis, with the diatoms nested within the Bigelowiella plus Planoglabratella (Rhizaria) grouping. The particular relationships recovered are further consistent with two characteristic sequence motifs. The best explanation of our data analyses is an EFL gene transfer from a foraminifer to a diatom, the first case in which the donor–recipient relationship was clarified. Finally, based on a reverse transcriptase quantitative PCR assay and the genome information of Thalassiosira and Phaeodactylum, we propose the loss of elongation factor function in Thalassiosira EF-1α. PMID:18458344

  2. Indication of Horizontal DNA Gene Transfer by Extracellular Vesicles

    PubMed Central

    Speiseder, Thomas; Badbaran, Anita; Reimer, Rudolph; Indenbirken, Daniela; Grundhoff, Adam; Brunswig-Spickenheier, Bärbel; Alawi, Malik; Lange, Claudia

    2016-01-01

    The biological relevance of extracellular vesicles (EV) in intercellular communication has been well established. Thus far, proteins and RNA were described as main cargo. Here, we show that EV released from human bone marrow derived mesenchymal stromal cells (BM-hMSC) also carry high-molecular DNA in addition. Extensive EV characterization revealed this DNA mainly associated with the outer EV membrane and to a smaller degree also inside the EV. Our EV purification protocol secured that DNA is not derived from apoptotic or necrotic cells. To analyze the relevance of EV-associated DNA we lentivirally transduced Arabidopsis thaliana-DNA (A.t.-DNA) as indicator into BM-hMSC and generated EV. Using quantitative polymerase chain reaction (qPCR) techniques we detected high copy numbers of A.t.-DNA in EV. In recipient hMSC incubated with tagged EV for two weeks we identified A.t.-DNA transferred to recipient cells. Investigation of recipient cell DNA using quantitative PCR and verification of PCR-products by sequencing suggested stable integration of A.t.-DNA. In conclusion, for the first time our proof-of-principle experiments point to horizontal DNA transfer into recipient cells via EV. Based on our results we assume that eukaryotic cells are able to exchange genetic information in form of DNA extending the known cargo of EV by genomic DNA. This mechanism might be of relevance in cancer but also during cell evolution and development. PMID:27684368

  3. Horizontal transfer of a large and highly toxic secondary metabolic gene cluster between fungi.

    PubMed

    Slot, Jason C; Rokas, Antonis

    2011-01-25

    Genes involved in intermediary and secondary metabolism in fungi are frequently physically linked or clustered. For example, in Aspergillus nidulans the entire pathway for the production of sterigmatocystin (ST), a highly toxic secondary metabolite and a precursor to the aflatoxins (AF), is located in a ∼54 kb, 23 gene cluster. We discovered that a complete ST gene cluster in Podospora anserina was horizontally transferred from Aspergillus. Phylogenetic analysis shows that most Podospora cluster genes are adjacent to or nested within Aspergillus cluster genes, although the two genera belong to different taxonomic classes. Furthermore, the Podospora cluster is highly conserved in content, sequence, and microsynteny with the Aspergillus ST/AF clusters and its intergenic regions contain 14 putative binding sites for AflR, the transcription factor required for activation of the ST/AF biosynthetic genes. Examination of ∼52,000 Podospora expressed sequence tags identified transcripts for 14 genes in the cluster, with several expressed at multiple life cycle stages. The presence of putative AflR-binding sites and the expression evidence for several cluster genes, coupled with the recent independent discovery of ST production in Podospora [1], suggest that this HGT event probably resulted in a functional cluster. Given the abundance of metabolic gene clusters in fungi, our finding that one of the largest known metabolic gene clusters moved intact between species suggests that such transfers might have significantly contributed to fungal metabolic diversity. PAPERFLICK:

  4. Adenovirus-mediated gene transfer to tumor cells.

    PubMed

    Cascalló, Manel; Alemany, Ramon

    2004-01-01

    Cell transduction in vitro is only the first step toward proving that a genetherapy vector can be useful to treat tumors. However, tumor targeting in vivo is now the milestone for gene therapy to succeed against disseminated cancer. Therefore, most valuable information is obtained from studies of vector biodistribution. Owing to the hepatotropism of adenoviral vectors, a particularly important parameter is the tumor/liver ratio. This ratio can be given at the level of gene expression if the amount of transgene expression is measured. To optimize the targeting, however, the levels of viral particles that reach the tumor compared to other organs must be studied. Most of this chapter deals with methods to quantify the virus fate in tumor-bearing animals. We present a radioactive labeling method that can be used to study biodistribution. After a small section dealing with tumor models, we describe methods to quantify different parameters related to adenovirus-mediated tumor targeting.

  5. Volcanic hazard warning system: Persistence and transferability

    NASA Astrophysics Data System (ADS)

    Sorensen, John H.; Gersmehl, Philip J.

    1980-03-01

    This study examines the normative functioning of the volcano warning system on the Island of Hawaii. The research seeks to identify the combination of factors, both environmental and social, which allows the system to operate with success. On the basis of the findings, several recommendations are offered for improving environmental hazard warning systems.

  6. Gene Transfer into Rat Brain Using Adenoviral Vectors

    PubMed Central

    Puntel, Mariana; Kroeger, Kurt M.; Sanderson, Nicholas S.R.; Thomas, Clare E.; Castro, Maria G.; Lowenstein, Pedro R.

    2010-01-01

    Viral vector–mediated gene delivery is an attractive procedure for introducing genes into the brain, both for purposes of basic neuroscience research and to develop gene therapy for neurological diseases. Replication-defective adenoviruses possess many features which make them ideal vectors for this purpose—efficiently transducing terminally differentiated cells such as neurons and glial cells, resulting in high levels of transgene expression in vivo. Also, in the absence of anti-adenovirus immunity, these vectors can sustain very long-term transgene expression within the brain parenchyma. This unit provides protocols for the stereotactic injection of adenoviral vectors into the brain, followed by protocols to detect transgene expression or infiltrates of immune cells by immunocytochemistry or immunofluorescence. ELISPOT and neutralizing antibody assay methodologies are provided to quantitate the levels of cellular and humoral immune responses against adenoviruses. Quantitation of adenoviral vector genomes within the rat brain using qPCR is also described. Curr. Protoc. Neurosci. 50:4.24.1–4.24.49. © 2010 by John Wiley & Sons, Inc. PMID:20066657

  7. Herpes simplex virus-mediated human hypoxanthine-guanine phosphoribosyltransferase gene transfer into neuronal cells

    SciTech Connect

    Palella, T.D.; Silverman, L.J.; Schroll, C.T.; Homa, F.L.; Levine, M.; Kelley, W.N.

    1988-01-01

    The virtually complete deficiency of the purine salvage enzyme hypoxanthine-guanine phosphoribosyltransferase (HPRT) results in a devastating neurological disease, Lesch-Nyhan syndrome. Transfer of the HPRT gene into fibroblasts and lymphoblasts in vitro and into hematopoietic cells in vivo has been accomplished by other groups with retroviral-derived vectors. It appears to be necessary, however, to transfer the HPRT gene into neuronal cells to correct the neurological dysfunction of this disorder. The neurotropic virus herpes simplex virus type 1 has features that make it suitable for use as a vector to transfer the HPRT gene into neuronal tissue. This report describes the isolation of an HPRT-deficient rat neuroma cell line, designated B103-4C, and the construction of a recombinant herpes simplex virus type 1 that contained human HPRT cDNA. These recombinant viruses were used to infect B103-4C cells. Infected cells expressed HPRT activity which was human in origin.

  8. Cloning of the organophosphorus pesticide hydrolase gene clusters of seven degradative bacteria isolated from a methyl parathion contaminated site and evidence of their horizontal gene transfer.

    PubMed

    Zhang, Ruifu; Cui, Zhongli; Zhang, Xiaozhou; Jiang, Jiandong; Gu, Ji-Dong; Li, Shunpeng

    2006-10-01

    Seven organophosphorus pesticide-degrading bacteria harboring the methyl parathion degrading (mpd) gene were isolated from a methyl parathion contaminated site. In this study, the 4.7 kb mpd gene cluster, conserved in all seven bacteria capable of degrading methyl parathion, was cloned and further analysis revealed that this cluster contained five ORFs and the mpd gene was associated with a mobile element, IS6100. In addition to mpd gene ORF and tnpA ORF, three other ORFs showed high homology to the permease component of ABC-type transport system, the general secretion pathway protein B, and the RNA polymerase sigma 70 factor, respectively. The mpd genes of these 7 strains were subcloned and expressed in E. coli, SDS-PAGE and zymogram analysis showed that two expression products of mpd genes in E. coli were found, but the one without signal peptide showed the hydrolytic activities. Our evidences collectively suggest that mpd gene cluster may be disseminated through horizontal gene transfer based on phylogenetic analysis of the cluster and their host bacterial strains, and comparisons of GC content of the cluster and respective host's chromosome.

  9. Antibody gene transfer with adeno-associated viral vectors as a method for HIV prevention.

    PubMed

    Brady, Jacqueline M; Baltimore, David; Balazs, Alejandro B

    2017-01-01

    Broadly neutralizing antibodies (bNAbs) against human immunodeficiency virus (HIV) show great promise in HIV prevention as they are capable of potently neutralizing a considerable breadth of genetically diverse strains. Passive transfer of monoclonal bNAb proteins can confer protection in animal models of HIV infection at modest concentrations, inspiring efforts to develop an HIV vaccine capable of eliciting bNAb responses. However, these antibodies demonstrate high degrees of somatic mutation and other unique characteristics that may hinder the ability of conventional approaches to consistently and effectively produce bNAb analogs. As an alternative strategy, we and others have proposed vector-mediated gene transfer to generate long-term, systemic production of bNAbs in the absence of immunization. Herein, we review the use of adeno-associated virus (AAV) vectors for delivery of HIV bNAbs and antibody-like proteins and summarize both the advantages and disadvantages of this strategy as a method for HIV prevention.

  10. Viral Vectors for In Vivo Gene Transfer in Parkinson’s disease: Properties and Clinical Grade Production

    PubMed Central

    Burger, Corinna; Snyder, Richard O.

    2009-01-01

    Because Parkinson’s disease is a progressive degenerative disorder that is mainly confined to the basal ganglia, gene transfer to deliver therapeutic molecules is an attractive treatment avenue. The present review focuses on direct in vivo gene transfer vectors that have been developed to a degree that they have been successfully used in animal model of Parkinson’s disease. Accordingly, the properties of recombinant adenovirus, recombinant adeno-associated virus, herpes simplex virus, and lentivirus are described and contrasted. In order for viral vectors to be developed into clinical grade reagents, they must be manufactured and tested to precise regulatory standards. Indeed, clinical lots of viral vectors can be produced in compliance with current Good Manufacturing Practices (cGMPs) regulations using industry accepted manufacturing methodologies, manufacturing controls, and quality systems. The viral vector properties themselves combined with physiological product formulations facilitate long-term storage and direct in vivo administration. PMID:17916354

  11. NIH oversight of human gene transfer research involving retroviral, lentiviral, and adeno-associated virus vectors and the role of the NIH recombinant DNA advisory committee.

    PubMed

    O'Reilly, Marina; Shipp, Allan; Rosenthal, Eugene; Jambou, Robert; Shih, Tom; Montgomery, Maureen; Gargiulo, Linda; Patterson, Amy; Corrigan-Curay, Jacqueline

    2012-01-01

    In response to public and scientific concerns regarding human gene transfer research, the National Institutes of Health (NIH) developed a transparent oversight system that extends to human gene transfer protocols that are either conducted with NIH funding or conducted at institutions that receive NIH funding for recombinant DNA research. The NIH Recombinant DNA Advisory Committee (RAC) has been the primary advisory body to NIH regarding the conduct of this research. Human gene transfer research proposals that are subject to the NIH Guidelines for Research Involving Recombinant DNA Molecules (NIH Guidelines) must be submitted to the NIH Office of Biotechnology Activities (OBA), and protocols that raise novel scientific, safety, medical, ethical, or social issues are publicly discussed at the RAC's quarterly public meetings. OBA also convenes gene transfer safety symposia and policy conferences to provide a public forum for scientific experts to discuss emerging issues in the field. This transparent system of review promotes the rapid exchange of important scientific information and dissemination of data. The goal is to optimize the conduct of individual research protocols and to advance gene transfer research generally. This process has fostered the development of retroviral, lentiviral, and adeno-associated viral vector mediated gene delivery.

  12. The National Institutes of Health Oversight of Human Gene Transfer Research: Enhancing Science and Safety.

    PubMed

    O'Reilly, Marina; Jambou, Robert; Rosenthal, Eugene; Montgomery, Maureen; Hassani, Morad; Gargiulo, Linda; Corrigan-Curay, Jacqueline

    2015-01-01

    The National Institutes of Health (NIH) oversight of human gene transfer research, which is defined as the deliberate transfer of recombinant and/or synthetic nucleic acid molecules to humans, originates with the NIH Guidelines for Research Involving Recombinant or Synthetic Nucleic Acid Molecules (NIH Guidelines). The NIH Guidelines, which were first published in the Federal Register almost 40 years ago, have been amended numerous times to remain responsive to scientific progress and to clearly define the responsibilities of NIH, the Recombinant DNA Advisory Committee (RAC), investigators, and institutions. Human gene transfer trials conducted at clinical sites in the United States (USA) are subject to the NIH Guidelines if they are conducted at, or sponsored by, an institution that receives any support for recombinant or synthetic nucleic acid research from the NIH. Human gene transfer trials conducted either in the USA or abroad are also subject to the NIH Guidelines if the investigational agent was developed with NIH funds and the institution that developed the investigational materials sponsors or participates in these projects. Trials are registered with the NIH Office Biotechnology Activities (OBA) and there are ongoing reporting requirements. Each new trial is reviewed by the RAC, and those that are novel or raise unique ethical or social issues are selected for review at quarterly public RAC meetings. The RAC also advises the NIH on policy and other matters relating to clinical gene transfer research and biosafety.

  13. An Efficient Low Cost Method for Gene Transfer to T Lymphocytes

    PubMed Central

    Chicaybam, Leonardo; Sodre, Andressa Laino; Curzio, Bianca Azevedo; Bonamino, Martin Hernan

    2013-01-01

    Gene transfer to T lymphocytes has historically relied on retro and lentivirus, but recently transposon-based gene transfer is rising as a simpler and straight forward approach to achieve stable transgene expression. Transfer of expression cassettes to T lymphocytes remains challenging, being based mainly on commercial kits. Aims We herein report a convenient and affordable method based on in house made buffers, generic cuvettes and utilization of the widely available Lonza nucleofector II device to promote efficient gene transfer to T lymphocytes. Results This approach renders high transgene expression levels in primary human T lymphocytes (mean 45%, 41–59%), the hard to transfect murine T cells (mean 38%, 36–42% for C57/BL6 strain) and human Jurkat T cell line. Cell viability levels after electroporation allowed further manipulations such as in vitro expansion and Chimeric Antigen Receptor (CAR) mediated gain of function for target cell lysis. Conclusions We describe here an efficient general protocol for electroporation based modification of T lymphocytes. By opening access to this protocol, we expect that efficient gene transfer to T lymphocytes, for transient or stable expression, may be achieved by an increased number of laboratories at lower and affordable costs. PMID:23555950

  14. Adenoviral-Mediated Imaging of Gene Transfer Using a Somatostatin Receptor-Cytosine Deaminase Fusion Protein

    PubMed Central

    Lears, Kimberly A.; Parry, Jesse J.; Andrews, Rebecca; Nguyen, Kim; Wadas, Thaddeus J.; Rogers, Buck E.

    2015-01-01

    Suicide gene therapy is a process by which cells are administered a gene that encodes a protein capable of converting a nontoxic prodrug into an active toxin. Cytosine deaminase (CD) has been widely investigated as a means of suicide gene therapy due to the enzyme’s ability to convert the prodrug 5-fluorocytosine (5-FC) into the toxic compound 5-fluorouracil (5-FU). However, the extent of gene transfer is a limiting factor in predicting therapeutic outcome. The ability to monitor gene transfer, non-invasively, would strengthen the efficiency of therapy. In this regard, we have constructed and evaluated a replication-deficient adenovirus (Ad) containing the human somatostatin receptor subtype 2 (SSTR2) fused with a C-terminal yeast CD gene for the non-invasive monitoring of gene transfer and therapy. The resulting Ad (AdSSTR2-yCD) was evaluated in vitro in breast cancer cells to determine the function of the fusion protein. These studies demonstrated that the both the SSTR2 and yCD were functional in binding assays, conversion assays, and cytotoxicity assays. In vivo studies similarly demonstrated the functionality using conversion assays, biodistribution studies, and small animal positron-emission tomography (PET) imaging studies. In conclusion, the fusion protein has been validated as useful for the non-invasive imaging of yCD expression and will be evaluated in the future for monitoring yCD-based therapy. PMID:25837665

  15. Adenoviral-mediated imaging of gene transfer using a somatostatin receptor-cytosine deaminase fusion protein.

    PubMed

    Lears, K A; Parry, J J; Andrews, R; Nguyen, K; Wadas, T J; Rogers, B E

    2015-03-01

    Suicide gene therapy is a process by which cells are administered a gene that encodes a protein capable of converting a nontoxic prodrug into an active toxin. Cytosine deaminase (CD) has been widely investigated as a means of suicide gene therapy owing to the enzyme's ability to convert the prodrug 5-fluorocytosine (5-FC) into the toxic compound 5-fluorouracil (5-FU). However, the extent of gene transfer is a limiting factor in predicting therapeutic outcome. The ability to monitor gene transfer, non-invasively, would strengthen the efficiency of therapy. In this regard, we have constructed and evaluated a replication-deficient adenovirus (Ad) containing the human somatostatin receptor subtype 2 (SSTR2) fused with a C-terminal yeast CD gene for the non-invasive monitoring of gene transfer and therapy. The resulting Ad (AdSSTR2-yCD) was evaluated in vitro in breast cancer cells to determine the function of the fusion protein. These studies demonstrated that both the SSTR2 and yCD were functional in binding assays, conversion assays and cytotoxicity assays. In vivo studies similarly demonstrated the functionality using conversion assays, biodistribution studies and small animal positron-emission tomography (PET) imaging studies. In conclusion, the fusion protein has been validated as useful for the non-invasive imaging of yCD expression and will be evaluated in the future for monitoring yCD-based therapy.

  16. Influence of earthworm activity on gene transfer from Pseudomonas fluorescens to indigenous soil bacteria.

    PubMed Central

    Daane, L L; Molina, J A; Berry, E C; Sadowsky, M J

    1996-01-01

    We have developed a model system to assess the influence of earthworm activity on the transfer of plasmid pJP4 from an inoculated donor bacterium, Pseudomonas fluorescens C5t (pJP4), to indigenous soil microorganisms. Three different earthworm species (Lumbricus terrestris, Lumbricus rubellus, and Aporrectodea trapezoides), each with unique burrowing, casting, and feeding behaviors, were evaluated. Soil columns were inoculated on the surface with 10(8) cells per g of soil of the donor bacterium, and after a 2-week incubation period, donor, transconjugant, and total bacteria were enumerated at 5-cm-depth intervals. Transconjugants were confirmed by use of colony hybridization with a mer gene probe. In situ gene transfer of plasmid pJP4 from P. fluorescens C5t to indigenous soil bacteria was detected in all inoculated microcosms. In the absence of earthworms, the depth of recovery was limited to the top 5 cm of the column, with approximately 10(3) transconjugants per g of soil. However, the total number of transconjugants recovered from soil was significantly greater in microcosms containing either L. rubellus or A. trapezoides, with levels reaching about 10(5) CFU/g of soil. In addition, earthworms distributed donor and transconjugant bacteria throughout the microcosm columns, with the depth of recovery dependent on the burrowing behavior of each earthworm species. Donor and transconjugant bacteria were also recovered from earthworm casts and inside developing cocoons. Transconjugant bacteria from the indigenous soil microflora were classified as belonging to Acidovorax spp., Acinetobacter spp., Agrobacterium spp., Pasteurella spp., Pseudomonas spp., and Xanthomonas spp. PMID:8593052

  17. Transferring Gus gene into intact rice cells by low energy ion beam

    NASA Astrophysics Data System (ADS)

    Zengliang, Yu; Jianbo, Yang; Yuejin, Wu; Beijiu, Cheng; Jianjun, He; Yuping, Huo

    1993-06-01

    A new technique of transferring genes by low energy ion beam has been reported in this paper. The Gus and CAT (chloramphenicol acetyltransferase) genes, as "foreign" genetic materials, were introduced into the suspension cells and ripe embryos or rice by implantation of 20-30 keV Ar + at doses ranging from 1 × 10 15 to 4 × 10 15 ions/cm 2. The activities of CAT and Gus were detected in the cells and embryos after several weeks. The results indicate that the transfer was a success.

  18. An Efficient Algorithm for Gene/Species Trees Parsimonious Reconciliation with Losses, Duplications and Transfers

    NASA Astrophysics Data System (ADS)

    Doyon, Jean-Philippe; Scornavacca, Celine; Gorbunov, K. Yu.; Szöllősi, Gergely J.; Ranwez, Vincent; Berry, Vincent

    Tree reconciliation methods aim at estimating the evolutionary events that cause discrepancy between gene trees and species trees. We provide a discrete computational model that considers duplications, transfers and losses of genes. The model yields a fast and exact algorithm to infer time consistent and most parsimonious reconciliations. Then we study the conditions under which parsimony is able to accurately infer such events. Overall, it performs well even under realistic rates, transfers being in general less accurately recovered than duplications. An implementation is freely available at http://www.atgc-montpellier.fr/MPR.

  19. Distant horizontal gene transfer is rare for multiple families of prokaryotic insertion sequences.

    PubMed

    Wagner, Andreas; de la Chaux, Nicole

    2008-11-01

    Horizontal gene transfer in prokaryotes is rampant on short and intermediate evolutionary time scales. It poses a fundamental problem to our ability to reconstruct the evolutionary tree of life. Is it also frequent over long evolutionary distances? To address this question, we analyzed the evolution of 2,091 insertion sequences from all 20 major families in 438 completely sequenced prokaryotic genomes. Specifically, we mapped insertion sequence occurrence on a 16S rDNA tree of the genomes we analyzed, and we also constructed phylogenetic trees of the insertion sequence transposase coding sequences. We found only 30 cases of likely horizontal transfer among distantly related prokaryotic clades. Most of these horizontal transfer events are ancient. Only seven events are recent. Almost all of these transfer events occur between pairs of human pathogens or commensals. If true also for other, non-mobile DNA, the rarity of distant horizontal transfer increases the odds of reliable phylogenetic inference from sequence data.

  20. The effect of mucolytic agents on gene transfer across a CF sputum barrier in vitro.

    PubMed

    Stern, M; Caplen, N J; Browning, J E; Griesenbach, U; Sorgi, F; Huang, L; Gruenert, D C; Marriot, C; Crystal, R G; Geddes, D M; Alton, E W

    1998-01-01

    Trials of gene transfer for cystic fibrosis (CF) are currently underway. However, direct application to the airways may be impeded by the presence of airway secretions. We have therefore assessed the effect of CF sputum on the expression of the reporter gene beta-galactosidase complexed with the cationic liposome DC-Chol/DOPE in a number of cell lines in vitro. Transfection was markedly inhibited in the presence of sputum; the effect was concentration dependent and was only partially ameliorated by removal of sputum with phosphate-buffered saline (PBS) washing before gene transfer. However, treatment of the sputum-covered cells with recombinant human DNase (rhDNase, 50 micrograms/ml) but not with N-acetylcysteine, Nacystelyn, lysine (all 20 mM) or recombinant alginase (0.5 U/ml) significantly (P < 0.005) improved gene transfer. Adenovirus-mediated gene transfer efficiency in the presence of sputum was similarly inhibited, and again, treatment with rhDNase before transfection significantly improved gene transfer (P < 0.005). Transfection of Cos 7 cells in the presence of exogenous genomic DNA alone demonstrated similar inhibition to that observed with sputum and was also ameliorated by pre-treatment of DNA-covered cells with rhDNase. In a separate series of experiments performed in the absence of added sputum or genomic DNA, increasing concentrations of rhDNase resulted in a concentration-related decline in transfection efficiency. However, even at the highest concentration (500 micrograms/ml of rhDNase), transfection efficiency remained more than 50% of control. Thus, pre-treatment of CF airways with rhDNase may be appropriate before liposome or adenovirus-mediated gene therapy.

  1. Plug-to-plug gas transfer system

    DOEpatents

    Poindexter, Allan M.

    1978-01-01

    A system for conducting a fluid from one component to another component of a nuclear reactor wherein at least one such component is a rotatable closure head plug capable of movement relative to the other component. The conducting system utilizes the annulus located between the components as a connecting passageway for the fluid.

  2. Regulatory and ethical issues for phase I in utero gene transfer studies.

    PubMed

    Strong, Carson

    2011-11-01

    Clinical gene transfer research has involved adult and child subjects, and it is expected that gene transfer in fetal subjects will occur in the future. Some genetic diseases have serious adverse effects on the fetus before birth, and there is hope that prenatal gene therapy could prevent such disease progression. Research in animal models of prenatal gene transfer is actively being pursued. The prospect of human phase I in utero gene transfer studies raises important regulatory and ethical issues. One issue not previously addressed arises in applying U.S. research regulations to such studies. Specifically, current regulations state that research involving greater than minimal risk to the fetus and no prospect of direct benefit to the fetus or pregnant woman is not permitted. Phase I studies will involve interventions such as needle insertions through the uterus, which carry risks to the fetus including spontaneous abortion and preterm birth. It is possible that these risks will be regarded as exceeding minimal. Also, some regard the probability of therapeutic benefit in phase I studies to be so low that these studies do not satisfy the regulatory requirement that they "hold out the prospect of direct benefit" to subjects. On the basis of these considerations, investigators and institutional review boards might reasonably conclude that some phase I in utero studies are not to be permitted. This paper identifies considerations that are relevant to such judgments and explores ethically acceptable ways in which phase I studies can be designed so that they are permitted by the regulations.

  3. Endosomal processing limits gene transfer to polarized airway epithelia by adeno-associated virus

    PubMed Central

    Duan, Dongsheng; Yue, Yongping; Yan, Ziying; Yang, Jusan; Engelhardt, John F.

    2000-01-01

    The restriction of viral receptors and coreceptors to the basolateral surface of airway epithelial cells has been blamed for the inefficient transfer of viral vectors to the apical surface of this tissue. We now report, however, that differentiated human airway epithelia internalize rAAV type-2 virus efficiently from their apical surfaces, despite the absence of known adeno-associated virus–2 (AAV-2) receptors or coreceptors at these sites. The dramatically lower transduction efficiency of rAAV infection from the apical surface of airway cells appears to result instead from differences in endosomal processing and nuclear trafficking of apically or basolaterally internalized virions. AAV capsid proteins are ubiquitinated after endocytosis, and gene transfer can be significantly enhanced by proteasome or ubiquitin ligase inhibitors. Tripeptide proteasome inhibitors increased persistent rAAV gene delivery from the apical surface >200-fold, to a level nearly equivalent to that achieved with basolateral infection. In vivo application of proteasome inhibitor in mouse lung augmented rAAV gene transfer from undetectable levels to a mean of 10.4 ± 1.6% of the epithelial cells in large bronchioles. Proteasome inhibitors also increased rAAV-2–mediated gene transfer to the liver tenfold, but they did not affect transduction of skeletal or cardiac muscle. These findings suggest that tissue-specific ubiquitination of viral capsid proteins interferes with rAAV-2 transduction and provides new approaches to circumvent this barrier for gene therapy of diseases such as cystic fibrosis. PMID:10841516

  4. Gene transfer to muscle from the isolated regional circulation.

    PubMed

    Petrov, Mihail; Malik, Alock; Mead, Andrew; Bridges, Charles R; Stedman, Hansell H

    2011-01-01

    Vector transport across the endothelium has long been regarded as one of the central "bottlenecks" in gene therapy research, especially as it pertains to the muscular dystrophies where the target tissue approaches half of the total body mass. Clinical studies of gene therapy for hemophilia B revealed the limitations of the intramuscular route, compelling an aggressive approach to the study of scale-independent circulatory means of vector delivery. The apparent permeability of the microvasculature in small animals suggests that gravitational and/or inertial effects on the circulation require progressive restriction of fluid and solute flow across the capillary wall with increasing body size. To overcome this physiological restriction, we initially used a combined surgical and pharmacological approach to temporarily alter permeability within the isolated pelvic limb. Although this was successful, new information about the cell and molecular biology of histamine-induced changes in microvascular permeability suggested an alternative approach, which substituted pressure-induced transvenular extravasation. Here we outline the details of our surgical approaches in the rat. We also discuss the modifications that are appropriate for the dog.

  5. Radiative heat transfer in low-dimensional systems -- microscopic mode

    NASA Astrophysics Data System (ADS)

    Woods, Lilia; Phan, Anh; Drosdoff, David

    2013-03-01

    Radiative heat transfer between objects can increase dramatically at sub-wavelength scales. Exploring ways to modulate such transport between nano-systems is a key issue from fundamental and applied points of view. We advance the theoretical understanding of radiative heat transfer between nano-objects by introducing a microscopic model, which takes into account the individual atoms and their atomic polarizabilities. This approach is especially useful to investigate nano-objects with various geometries and give a detailed description of the heat transfer distribution. We employ this model to study the heat exchange in graphene nanoribbon/substrate systems. Our results for the distance separations, substrates, and presence of extended or localized defects enable making predictions for tailoring the radiative heat transfer at the nanoscale. Financial support from the Department of Energy under Contract No. DE-FG02-06ER46297 is acknowledged.

  6. Collective evolution of cyanobacteria and cyanophages mediated by horizontal gene transfer

    NASA Astrophysics Data System (ADS)

    Shih, Hong-Yan; Rogers, Tim; Goldenfeld, Nigel

    We describe a model for how antagonistic predator-prey coevolution can lead to mutualistic adaptation to an environment, as a result of horizontal gene transfer. Our model is a simple description of ecosystems such as marine cyanobacteria and their predator cyanophages, which carry photosynthesis genes. These genes evolve more rapidly in the virosphere than the bacterial pan-genome, and thus the bacterial population could potentially benefit from phage predation. By modeling both the barrier to predation and horizontal gene transfer, we study this balance between individual sacrifice and collective benefits. The outcome is an emergent mutualistic coevolution of improved photosynthesis capability, benefiting both bacteria and phage. This form of multi-level selection can contribute to niche stratification in the cyanobacteria-phage ecosystem. This work is supported in part by a cooperative agreement with NASA, Grant NNA13AA91A/A0018.

  7. Endosperm transfer cell-specific genes and proteins: structure, function and applications in biotechnology

    PubMed Central

    Lopato, Sergiy; Borisjuk, Nikolai; Langridge, Peter; Hrmova, Maria

    2014-01-01

    Endosperm transfer cells (ETC) are one of four main types of cells in endosperm. A characteristic feature of ETC is the presence of cell wall in-growths that create an enlarged plasma membrane surface area. This specialized cell structure is important for the specific function of ETC, which is to transfer nutrients from maternal vascular tissue to endosperm. ETC-specific genes are of particular interest to plant biotechnologists, who use genetic engineering to improve grain quality and yield characteristics of important field crops. The success of molecular biology-based approaches to manipulating ETC function is dependent on a thorough understanding of the functions of ETC-specific genes and ETC-specific promoters. The aim of this review is to summarize the existing data on structure and function of ETC-specific genes and their products. Potential applications of ETC-specific genes, and in particular their promoters for biotechnology will be discussed. PMID:24578704

  8. Fishing for biodiversity: novel methanopterin-linked C transfer genes deduced from the Sargasso Sea metagenome.

    PubMed

    Kalyuzhnaya, Marina G; Nercessian, Olivier; Lapidus, Alla; Chistoserdova, Ludmila

    2005-12-01

    The recently generated database of microbial genes from an oligotrophic environment populated by a calculated 1800 major phylotypes (the Sargasso Sea metagenome-SSM) presents a great source for expanding local databases of genes indicative of a specific function. In this article we analyse the SSM for the presence of methanopterin-linked C1 transfer genes that are signature for methylotrophy. We conclude that more than 10 phylotypes possessing genes of interest are present in this environment. The sequences representative of these major phylotypes do not appear to belong to any known microbial group capable of methanopterin-linked C1 transfer. Instead, these sequences separate from all known sequences on phylogenetic trees, pointing toward their affiliation with novel microbial phyla. These data imply a broader distribution of methanopterin-linked functions in the microbial world than has been previously known.

  9. Origin of the plant Tm-1-like gene via two independent horizontal transfer events and one gene fusion event

    PubMed Central

    Yang, Zefeng; Liu, Li; Fang, Huimin; Li, Pengcheng; Xu, Shuhui; Cao, Wei; Xu, Chenwu; Huang, Jinling; Zhou, Yong

    2016-01-01

    The Tomato mosaic virus (ToMV) resistance gene Tm-1 encodes a direct inhibitor of ToMV RNA replication to protect tomato from infection. The plant Tm-1-like (Tm-1L) protein is predicted to contain an uncharacterized N-terminal UPF0261 domain and a C-terminal TIM-barrel signal transduction (TBST) domain. Homologous searches revealed that proteins containing both of these two domains are mainly present in charophyte green algae and land plants but absent from glaucophytes, red algae and chlorophyte green algae. Although Tm-1 homologs are widely present in bacteria, archaea and fungi, UPF0261- and TBST-domain-containing proteins are generally encoded by different genes in these linages. A co-evolution analysis also suggested a putative interaction between UPF0261- and TBST-domain-containing proteins. Phylogenetic analyses based on homologs of these two domains revealed that plants have acquired UPF0261- and TBST-domain-encoding genes through two independent horizontal gene transfer (HGT) events before the origin of land plants from charophytes. Subsequently, gene fusion occurred between these two horizontally acquired genes and resulted in the origin of the Tm-1L gene in streptophytes. Our results demonstrate a novel evolutionary mechanism through which the recipient organism may acquire genes with functional interaction through two different HGT events and further fuse them into one functional gene. PMID:27647002

  10. Single cell genomics indicates horizontal gene transfer and viral infections in a deep subsurface Firmicutes population

    PubMed Central

    Labonté, Jessica M.; Field, Erin K.; Lau, Maggie; Chivian, Dylan; Van Heerden, Esta; Wommack, K. Eric; Kieft, Thomas L.; Onstott, Tullis C.; Stepanauskas, Ramunas

    2015-01-01

    A major fraction of Earth's prokaryotic biomass dwells in the deep subsurface, where cellular abundances per volume of sample are lower, metabolism is slower, and generation times are longer than those in surface terrestrial and marine environments. How these conditions impact biotic interactions and evolutionary processes is largely unknown. Here we employed single cell genomics to analyze cell-to-cell genome content variability and signatures of horizontal gene transfer (HGT) and viral infections in five cells of Candidatus Desulforudis audaxviator, which were collected from a 3 km-deep fracture water in the 2.9 Ga-old Witwatersrand Basin of South Africa. Between 0 and 32% of genes recovered from single cells were not present in the original, metagenomic assembly of Desulforudis, which was obtained from a neighboring subsurface fracture. We found a transposable prophage, a retron, multiple clustered regularly interspaced short palindromic repeats (CRISPRs) and restriction-modification systems, and an unusually high frequency of transposases in the analyzed single cell genomes. This indicates that recombination, HGT and viral infections are prevalent evolutionary events in the studied population of microorganisms inhabiting a highly stable deep subsurface environment. PMID:25954269

  11. Construction of a Helicobacter pylori-Escherichia coli shuttle vector for gene transfer in Helicobacter pylori.

    PubMed Central

    Lee, W K; An, Y S; Kim, K H; Kim, S H; Song, J Y; Ryu, B D; Choi, Y J; Yoon, Y H; Baik, S C; Rhee, K H; Cho, M J

    1997-01-01

    In this study, a Helicobacter pylori-Escherichia coli shuttle vector was constructed for transferring DNA into H. pylori. The smallest cryptic plasmid (1.2 kb), pHP489, among those harbored by 77 H. pylori isolates was selected as a base replicon for constructing vectors. HindIII-digested pHP489 was ligated with a kanamycin resistance gene [aph(3')-III], which originated from Campylobacter jejuni, to produce the recombinant plasmid pHP489K. pHP489K was efficiently transformed into and stably maintained in H. pylori strains. The shuttle vector pBHP489K (3.6 kb) was constructed by the recombination of pHP489, ColE1, and aph(3')-III sequences. pBHP489K was reciprocally transformed into and maintained in both H. pylori and E. coli. Introduction of the shuttle vector clone DNA (pBHP489K/AB; 6.7 kb), containing the ureA and ureB genes of H. pylori, into urease-negative mutants of H. pylori led to the restoration of their urease activity. The transformants were confirmed to contain the incoming plasmid DNA. pBHP489K satisfied the requirements for an H. pylori-E. coli shuttle vector, implying that it might be a useful vector for investigating pathogenicity and restriction-modification systems of H. pylori. PMID:9406406

  12. Genome of Acanthamoeba castellanii highlights extensive lateral gene transfer and early evolution of tyrosine kinase signaling

    PubMed Central

    2013-01-01

    Background The Amoebozoa constitute one of the primary divisions of eukaryotes, encompassing taxa of both biomedical and evolutionary importance, yet its genomic diversity remains largely unsampled. Here we present an analysis of a whole genome assembly of Acanthamoeba castellanii (Ac) the first representative from a solitary free-living amoebozoan. Results Ac encodes 15,455 compact intron-rich genes, a significant number of which are predicted to have arisen through inter-kingdom lateral gene transfer (LGT). A majority of the LGT candidates have undergone a substantial degree of intronization and Ac appears to have incorporated them into established transcriptional programs. Ac manifests a complex signaling and cell communication repertoire, including a complete tyrosine kinase signaling toolkit and a comparable diversity of predicted extracellular receptors to that found in the facultatively multicellular dictyostelids. An important environmental host of a diverse range of bacteria and viruses, Ac utilizes a diverse repertoire of predicted pattern recognition receptors, many with predicted orthologous functions in the innate immune systems of higher organisms. Conclusions Our analysis highlights the important role of LGT in the biology of Ac and in the diversification of microbial eukaryotes. The early evolution of a key signaling facility implicated in the evolution of metazoan multicellularity strongly argues for its emergence early in the Unikont lineage. Overall, the availability of an Ac genome should aid in deciphering the biology of the Amoebozoa and facilitate functional genomic studies in this important model organism and environmental host. PMID:23375108

  13. Hepatic gene transfer in neonatal mice by adeno-associated virus serotype 8 vector.

    PubMed

    Wang, Lili; Wang, Huan; Bell, Peter; McMenamin, Deirdre; Wilson, James M

    2012-05-01

    For genetic diseases that manifest at a young age with irreversible consequences, early treatment is critical and essential. Neonatal gene therapy has the advantages of achieving therapeutic effects before disease manifestation, a low vector requirement and high vector-to-cell ratio, and a relatively immature immune system. Therapeutic effects or long-term rescue of neonatal lethality have been demonstrated in several animal models. However, vigorous cell proliferation in the newborn stage is a significant challenge for nonintegrating vectors, such as adeno-associated viral (AAV) vector. Slightly delaying the injection age, and readministration at a later time, are two of the alternative strategies to solve this problem. In this study, we demonstrated robust and efficient hepatic gene transfer by self-complementary AAV8 vector in neonatal mice. However, transduction quickly decreased over a few weeks because of vector dilution caused by fast proliferation. Delaying the injection age improved sustained expression, although it also increased neutralizing antibody (NAb) responses to AAV capsid. This approach can be used to treat genetic diseases with slow progression. For genetic diseases with early onset and severe consequences, early treatment is essential. A second injection of vector of a different serotype at a later time may overcome preexisting NAb and achieve sustained therapeutic effects.

  14. Gene Transfer from Targeted Liposomes to Specific Lymphoid Cells by Electroporation

    NASA Astrophysics Data System (ADS)

    Machy, Patrick; Lewis, Florence; McMillan, Lynette; Jonak, Zdenka L.

    1988-11-01

    Large unilamellar liposomes, coated with protein A and encapsulating the gene that confers resistance to mycophenolic acid, were used as a model system to demonstrate gene transfer into specific lymphoid cells. Protein A, which selectively recognizes mouse IgG2a antibodies, was coupled to liposomes to target them specifically to defined cell types coated with IgG2a antibody. Protein A-coated liposomes bound human B lymphoblastoid cells preincubated with a mouse IgG2a anti-HLA monoclonal antibody but failed to adhere to cells challenged with an irrelevant (anti-H-2) antibody of the same isotype or to cells incubated in the absence of antibody. Transfection of target cells bound to protein A-coated liposomes was achieved by electroporation. This step was essential since only electroporated cells survived in a selective medium containing mycophenolic acid. Transfection efficiency with electroporation and targeted liposomes was as efficient as conventional procedures that used unencapsulated plasmids free in solution but, in the latter case, cell selectivity is not possible. This technique provides a methodology for introducing defined biological macromolecules into specific cell types.

  15. Engineered Resilient Systems: Knowledge Capture and Transfer

    DTIC Science & Technology

    2014-08-29

    lly those made by semiconductor manufacturers , in order to achieve drastic reductions in schedule and cost of development for complex DoD-class...flexibility” in a system. Robust design has been traditionally applied to the manufacturing of systems, but also increasingly upstream to the concept design... additional interest in involving the AF/A9 organization in incorporating TSE-based approaches as they are responsible for integrating and coordinating

  16. Enhanced heat transfer in partially-saturated hydrothermal systems

    SciTech Connect

    Bixler, N.E.; Carrigan, C.R.

    1986-01-01

    The role of capillarity is potentially important for determining heat transfer in hydrothermal regions. Capillarity allows mixing of phases in liquid/vapor systems and results in enhanced two-phase convection. Comparisons involving a numerical model with capillarity and analytical models without indicate that heat transfer can be enhanced by about an order of magnitude. Whether capillarity can be important for a particular hydrothermal region will depend on the nature of mineral precipitation as well as pore and fracture size distributions.

  17. Highly efficient gene knockout by injection of TALEN mRNAs into oocytes and host transfer in Xenopus laevis.

    PubMed

    Nakajima, Keisuke; Yaoita, Yoshio

    2015-01-16

    Zinc-finger nucleases, transcription activator-like effector nucleases (TALENs) and the CRISPR/Cas (clustered regularly interspaced short palindromic repeats/CRISPR-associated proteins) system are potentially powerful tools for producing tailor-made knockout animals. However, their mutagenic activity is not high enough to induce mutations at all loci of a target gene throughout an entire tadpole. In this study, we present a highly efficient method for introducing gene modifications at almost all target sequences in randomly selected embryos. The gene modification activity of TALEN is enhanced by adopting the host-transfer technique. In our method, the efficiency is further improved by injecting TALEN mRNAs fused to the 3'UTR of the Xenopus DEADSouth gene into oocytes, which are then transferred into a host female frog, where they are ovulated and fertilized. The addition of the 3'UTR of the DEADSouth gene promotes mRNA translation in the oocytes and increases the expression of TALEN proteins to near-maximal levels three hours post fertilization (hpf). In contrast, TALEN mRNAs without this 3'UTR are translated infrequently in oocytes. Our data suggest that genomic DNA is more sensitive to TALEN proteins from fertilization to the midblastula (MBT) stage. Our method works by increasing the levels of TALEN proteins during the pre-MBT stages.

  18. Systematic Search for Evidence of Interdomain Horizontal Gene Transfer from Prokaryotes to Oomycete Lineages

    PubMed Central

    McCarthy, Charley G. P.

    2016-01-01

    ABSTRACT While most commonly associated with prokaryotes, horizontal gene transfer (HGT) can also have a significant influence on the evolution of microscopic eukaryotes. Systematic analysis of HGT in the genomes of the oomycetes, filamentous eukaryotic microorganisms in the Stramenopiles-Alveolates-Rhizaria (SAR) supergroup, has to date focused mainly on intradomain transfer events between oomycetes and fungi. Using systematic whole-genome analysis followed by phylogenetic reconstruction, we have investigated the extent of interdomain HGT between bacteria and plant-pathogenic oomycetes. We report five putative instances of HGT from bacteria into the oomycetes. Two transfers were found in Phytophthora species, including one unique to the cucurbit pathogen Phytophthora capsici. Two were found in Pythium species only, and the final transfer event was present in Phytopythium and Pythium species, the first reported bacterium-inherited genes in these genera. Our putative transfers included one protein that appears to be a member of the Pythium secretome, metabolic proteins, and enzymes that could potentially break down xenobiotics within the cell. Our findings complement both previous reports of bacterial genes in oomycete and SAR genomes and the growing body of evidence suggesting that interdomain transfer from prokaryotes into eukaryotes occurs more frequently than previously thought. IMPORTANCE Horizontal gene transfer (HGT) is the nonvertical inheritance of genetic material by transfer between different species. HGT is an important evolutionary mechanism for prokaryotes and in some cases is responsible for the spread of antibiotic resistance from resistant to benign species. Genome analysis has shown that examples of HGT are not as frequent in eukaryotes, but when they do occur they may have important evolutionary consequences. For example, the acquisition of fungal genes by an ancestral Phytophthora (plant destroyer) species is responsible for the large repertoire

  19. Learning Transfer--Validation of the Learning Transfer System Inventory in Portugal

    ERIC Educational Resources Information Center

    Velada, Raquel; Caetano, Antonio; Bates, Reid; Holton, Ed

    2009-01-01

    Purpose: The purpose of this paper is to analyze the construct validity of learning transfer system inventory (LTSI) for use in Portugal. Furthermore, it also aims to analyze whether LTSI dimensions differ across individual variables such as gender, age, educational level and job tenure. Design/methodology/approach: After a rigorous translation…

  20. Immunology of AAV-Mediated Gene Transfer in the Eye

    PubMed Central

    Willett, Keirnan; Bennett, Jean

    2013-01-01

    The eye has been at the forefront of translational gene therapy largely owing to suitable disease targets, anatomic accessibility, and well-studied immunologic privilege. These advantages have fostered research culminating in several clinical trials and adeno-associated virus (AAV) has emerged as the vector of choice for many ocular therapies. Pre-clinical and clinical investigations have assessed the humoral and cellular immune responses to a variety of naturally occurring and engineered AAV serotypes as well as their delivered transgenes and these data have been correlated to potential clinical sequelae. Encouragingly, AAV appears safe and effective with clinical follow-up surpassing 5 years in some studies. As disease targets continue to expand for AAV in the eye, thorough and deliberate assessment of immunologic safety is critical. With careful study, the development of these technologies should concurrently inform the biology of the ocular immune response. PMID:24009613

  1. Gene Transfer and the Reconstruction of Life's Early History from Genomic Data

    NASA Astrophysics Data System (ADS)

    Gogarten, J. Peter; Fournier, Gregory; Zhaxybayeva, Olga

    2008-03-01

    The metaphor of the unique and strictly bifurcating tree of life, suggested by Charles Darwin, needs to be replaced (or at least amended) to reflect and include processes that lead to the merging of and communication between independent lines of descent. Gene histories include and reflect processes such as gene transfer, symbioses and lineage fusion. No single molecule can serve as a proxy for the tree of life. Individual gene histories can be reconstructed from the growing molecular databases containing sequence and structural information. With some simplifications these gene histories can be represented by furcating trees; however, merging these gene histories into web-like organismal histories, including the transfer of metabolic pathways and cell biological innovations from now-extinct lineages, has yet to be accomplished. Because of these difficulties in interpreting the record retained in molecular sequences, correlations with biochemical fossils and with the geological record need to be interpreted with caution. Advances to detect and pinpoint transfer events promise to untangle at least a few of the intertwined histories of individual genes within organisms and trace them to the organismal ancestors. Furthermore, analysis of the shape of molecular phylogenetic trees may point towards organismal radiations that might reflect early mass extinction events that occurred on a planetary scale.

  2. Gene Transfer and the Reconstruction of Life's Early History from Genomic Data

    NASA Astrophysics Data System (ADS)

    Gogarten, J. Peter; Fournier, Gregory; Zhaxybayeva, Olga

    The metaphor of the unique and strictly bifurcating tree of life, suggested by Charles Darwin, needs to be replaced (or at least amended) to reflect and include processes that lead to the merging of and communication between independent lines of descent. Gene histories include and reflect processes such as gene transfer, symbioses and lineage fusion. No single molecule can serve as a proxy for the tree of life. Individual gene histories can be reconstructed from the growing molecular databases containing sequence and structural information. With some simplifications these gene histories can be represented by furcating trees; however, merging these gene histories into web-like organismal histories, including the transfer of metabolic pathways and cell biological innovations from now-extinct lineages, has yet to be accomplished. Because of these difficulties in interpreting the record retained in molecular sequences, correlations with biochemical fossils and with the geological record need to be interpreted with caution. Advances to detect and pinpoint transfer events promise to untangle at least a few of the intertwined histories of individual genes within organisms and trace them to the organismal ancestors. Furthermore, analysis of the shape of molecular phylogenetic trees may point towards organismal radiations that might reflect early mass extinction events that occurred on a planetary scale.

  3. Preventing High Fat Diet-induced Obesity and Improving Insulin Sensitivity through Neuregulin 4 Gene Transfer

    PubMed Central

    Ma, Yongjie; Gao, Mingming; Liu, Dexi

    2016-01-01

    Neuregulin 4 (NRG4), an epidermal growth factor-like signaling molecule, plays an important role in cell-to-cell communication during tissue development. Its function to regulate energy metabolism has recently been reported. This current study was designed to assess the preventive and therapeutic effects of NRG4 overexpression on high fat diet (HFD)-induced obesity. Using the hydrodynamic gene transfer method, we demonstrate that Nrg4 gene transfer in mice suppressed the development of diet-induced obesity, but did not affect pre-existing adiposity and body weight in obese mice. Nrg4 gene transfer curbed HFD-induced hepatic steatosis by inhibiting lipogenesis and PPARγ-mediated lipid storage. Concurrently, overexpression of NRG4 reduced chronic inflammation in both preventive and treatment studies, evidenced by lower mRNA levels of macrophage marker genes including F4/80, Cd68, Cd11b, Cd11c, and macrophage chemokine Mcp1, resulting in improved insulin sensitivity. Collectively, these results demonstrate that overexpression of the Nrg4 gene by hydrodynamic gene delivery prevents HFD-induced weight gain and fatty liver, alleviates obesity-induced chronic inflammation and insulin resistance, and supports the health benefits of NRG4 in managing obesity and obesity-associated metabolic disorders. PMID:27184920

  4. Fluoroquinolone induction of phage-mediated gene transfer in multidrug-resistant Salmonella.

    PubMed

    Bearson, Bradley L; Brunelle, Brian W

    2015-08-01

    Fluoroquinolones are broad-spectrum antibiotics that inhibit bacterial DNA gyrase and topoisomerase activity, which can cause DNA damage and result in bacterial cell death. In response to DNA damage, bacteria induce an SOS response to stimulate DNA repair. However, the SOS response may also induce prophage with production of infectious virions. Salmonella strains typically contain multiple prophages, and certain strains including phage types DT120 and DT104 contain prophage that upon induction are capable of generalised transduction. In this study, strains of multidrug-resistant (MDR) Salmonella enterica serovar Typhimurium DT120 and DT104 were exposed to fluoroquinolones important for use in human and veterinary disease therapy to determine whether prophage(s) are induced that could facilitate phage-mediated gene transfer. Cultures of MDR S. Typhimurium DT120 and DT104 containing a kanamycin resistance plasmid were lysed after exposure to fluoroquinolones (ciprofloxacin, enrofloxacin and danofloxacin). Bacterial cell lysates were able to transfer the plasmid to a recipient kanamycin-susceptible Salmonella strain by generalised transduction. In addition, exposure of DT120 to ciprofloxacin induced the recA gene of the bacterial SOS response and genes encoded in a P22-like generalised transducing prophage. This research indicates that fluoroquinolone exposure of MDR Salmonella can facilitate horizontal gene transfer, suggesting that fluoroquinolone usage in human and veterinary medicine may have unintended consequences, including the induction of phage-mediated gene transfer from MDR Salmonella. Stimulation of gene transfer following bacterial exposure to fluoroquinolones should be considered an adverse effect, and clinical decisions regarding antibiotic selection for infectious disease therapy should include this potential risk.

  5. Control System and Flexible Satellite Interaction During Orbit Transfer Maneuver

    NASA Technical Reports Server (NTRS)

    daSilva, Adenilson Roberto; GadelhadeSouza, Luiz Carlos

    1998-01-01

    In this paper the interaction between the attitude control system and the flexible structure of an artificial satellite during orbit transfer maneuver has been investigated. The satellite was modeled by a rigid central body with one or more flexible appendages. The dynamics equations were obtained by Lagrangean approach. The flexible appendages were treated as clamped-free beam and its displacement was discretized by assumed- mode method. In order to transfer the satellite, a typical Hohmann transfer and a burn-coast-burn strategy were used and the attitude was controlled by an on-off controller. During transfer procedure a global analysis of satellite has been done, such as: performance of control system, influence of elastic response in control system, thruster firing frequency, fuel consumption and variation of orbital elements. In order to avoid the interaction with structure motion, a control system with bandwidth of one decade bellow the fundamental frequency was used. In the simulations the firing frequency was evaluated in an approximately way but kept below the fundamental frequency of the structure. The control system has kept the attitude below the specifications. As a result, the orbit transfer maneuvering has been done correctly without excessive excitation of flexible appendage.

  6. Transfer Effects in Learning a Second Language Grammatical Gender System

    ERIC Educational Resources Information Center

    Sabourin, Laura; Stowe, Laurie A.; de Haan, Ger J.

    2006-01-01

    In this article second language (L2) knowledge of Dutch grammatical gender is investigated. Adult speakers of German, English and a Romance language (French, Italian or Spanish) were investigated to explore the role of transfer in learning the Dutch grammatical gender system. In the first language (L1) systems, German is the most similar to Dutch…

  7. Operational test report for 2706-T complex liquid transfer system

    SciTech Connect

    BENZEL, H.R.

    1999-09-01

    This document is the Operational Test Report (OTR). It enters the Record Copy of the W-259 Operational Test Procedure (HNF-3610) into the document retrieval system. Additionally, the OTR summarizes significant issues associated with testing the 2706-T waste liquid transfer and storage system.

  8. A Brief History of the Transfer System in British Columbia

    ERIC Educational Resources Information Center

    Gaber, Devron

    2005-01-01

    This document presents a brief history of important events in the development and maintenance of the post-secondary transfer system in British Columbia (BC) over the last six decades. Information is presented in point form and in chronological order. Several points related to the expansion of the post-secondary system are included, and hence the…

  9. 5 CFR 843.210 - Transfers between retirement systems.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 5 Administrative Personnel 2 2011-01-01 2011-01-01 false Transfers between retirement systems. 843.210 Section 843.210 Administrative Personnel OFFICE OF PERSONNEL MANAGEMENT (CONTINUED) CIVIL SERVICE REGULATIONS (CONTINUED) FEDERAL EMPLOYEES RETIREMENT SYSTEM-DEATH BENEFITS AND EMPLOYEE REFUNDS...

  10. 5 CFR 843.210 - Transfers between retirement systems.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 5 Administrative Personnel 2 2010-01-01 2010-01-01 false Transfers between retirement systems. 843.210 Section 843.210 Administrative Personnel OFFICE OF PERSONNEL MANAGEMENT (CONTINUED) CIVIL SERVICE REGULATIONS (CONTINUED) FEDERAL EMPLOYEES RETIREMENT SYSTEM-DEATH BENEFITS AND EMPLOYEE REFUNDS...

  11. Study of Lateral Gene Transfer in an Acid Mine Drainage Community Enabled by Comparative Genomics

    NASA Astrophysics Data System (ADS)

    Hugenholtz, P.; Croft, L.; Tyson, G. W.; Baker, B. J.; Detter, C.; Richardson, P. M.; Banfield, J. F.

    2002-12-01

    Lateral gene transfer (LGT) is thought to play a crucial role in the ecology and evolution of prokaryotes. We are investigating the role of LGT in an acid mine drainage community hosted in a pyrite-dominated metal sulfide deposit at the Richmond mine at Iron Mountain, CA. Due to biologically-mediated pyrite dissolution, the prevailing conditions within the mine are extremely low pH (< 1.0), very high ionic concentrations (molar concentrations of iron sulfate and mM concentrations of arsenic, copper and zinc), and moderate to high temperatures (30 to >50 C). These conditions are thought to largely isolate the community from potential external gene donors since naked DNA, phage and prokaryotes native to neutral pH habitats do not persist at pH <1.0 precluding an external influx of genes by transformation, transduction and conjugation, respectively. Microbial communities exist in several distinct habitats within Richmond mine including biofilms (subaqueous slime streamers and subaerial slimes) and cells attached directly to pyrite granules. This, however, belies an unusual simplicity in community composition. All communities investigated to date comprise only a handful of phylogenetically distinct organisms, typically dominated by the iron-oxidizing genera Leptospirillum and Ferroplasma. We have undertaken a community genomics analysis of a subaerial biofilm dominated by a Leptospirillum population to facilitate the study of LGT in this type of environment. The genome of Ferroplasma acidarmanus fer1, a minor component of the target community (but a major component of other Richmond mine communities), has been sequenced. Comparative genome analyses indicate that F. acidarmanus and the ancestor of two acidophilic Thermoplasma species belonging to the Euryarchaeota have traded many genes with phylogenetically remote acidophilic Sulfolobus species (Crenarchaeota). The putatively transferred sets of Sulfolobus genes in Ferroplasma and the Thermoplasma ancestor are distinct

  12. A case of horizontal gene transfer from Wolbachia to Aedes albopictus C6/36 cell line

    PubMed Central

    Hou, Qing; He, Ji; Yu, Jing; Ye, Yuting; Zhou, Dan; Sun, Yan; Zhang, Donghui; Ma, Lei; Shen, Bo; Zhu, Changliang

    2014-01-01

    Horizontal gene transfer plays an essential role in evolution and ecological adaptation, yet this phenomenon has remained controversial, particularly where it occurs between prokaryotes and eukaryotes. There are a handful of reported examples of horizontal gene transfer occurring between prokaryotes and eukaryotes in the literature, with most of these documented cases pertaining to invertebrates and endosymbionts. However, the vast majority of these horizontally transferred genes were either eventually excluded or rapidly became nonfunctional in the recipient genome. In this study, we report the discovery of a horizontal gene transfer from the endosymbiont Wolbachia in the C6/36 cell line derived from the mosquito Aedes albopictus. Moreover, we report that this horizontally transferred gene displayed high transcription level. This finding and the results of further experimentation strongly suggest this gene is functional and has been expressed and translated into a protein in the mosquito host cells. PMID:24812591

  13. Operon Formation is Driven by Co-Regulation and Not by Horizontal Gene Transfer

    SciTech Connect

    Price, Morgan N.; Huang, Katherine H.; Arkin, Adam P.; Alm, Eric J.

    2005-04-12

    Although operons are often subject to horizontal gene transfer (HGT), non-HGT genes are particularly likely to be in operons. To resolve this apparent discrepancy and to determine whether HGT is involved in operon formation, we examined the evolutionary history of the genes and operons in Escherichia coli K12. We show that genes that have homologs in distantly related bacteria but not in close relatives of E. coli (indicating HGTi) form new operons at about the same rates as native genes. Furthermore, genes in new operons are no more likely than other genes to have phylogenetic trees that are inconsistent with the species tree. In contrast, essential genes and ubiquitous genes without paralogs (genes believed to undergo HGT rarely) often form new operons. We conclude that HGT is not associated with operon formation, but instead promotes the prevalence of pre-existing operons. To explain operon formation, we propose that new operons reduce the amount of regulatory information required to specify optimal expression patterns. Consistent with this hypothesis, operons have greater amounts of conserved regulatory sequences than do individually transcribed genes.

  14. Role of horizontal gene transfer as a control on the coevolution of ribosomal proteins and the genetic code

    SciTech Connect

    Woese, Carl R.; Goldenfeld, Nigel; Luthey-Schulten, Zaida

    2011-03-31

    Our main goal is to develop the conceptual and computational tools necessary to understand the evolution of the universal processes of translation and replication and to identify events of horizontal gene transfer that occurred within the components. We will attempt to uncover the major evolutionary transitions that accompanied the development of protein synthesis by the ribosome and associated components of the translation apparatus. Our project goes beyond standard genomic approaches to explore homologs that are represented at both the structure and sequence level. Accordingly, use of structural phylogenetic analysis allows us to probe further back into deep evolutionary time than competing approaches, permitting greater resolution of primitive folds and structures. Specifically, our work focuses on the elements of translation, ranging from the emergence of the canonical genetic code to the evolution of specific protein folds, mediated by the predominance of horizontal gene transfer in early life. A unique element of this study is the explicit accounting for the impact of phenotype selection on translation, through a coevolutionary control mechanism. Our work contributes to DOE mission objectives through: (1) sophisticated computer simulation of protein dynamics and evolution, and the further refinement of techniques for structural phylogeny, which complement sequence information, leading to improved annotation of genomic databases; (2) development of evolutionary approaches to exploring cellular function and machinery in an integrated way; and (3) documentation of the phenotype interaction with translation over evolutionary time, reflecting the system response to changing selection pressures through horizontal gene transfer.

  15. Evolution of Acetoclastic Methanogenesis in Methanosarcina via Horizontal Gene Transfer from Cellulolytic Clostridia▿ †

    PubMed Central

    Fournier, Gregory P.; Gogarten, J. Peter

    2008-01-01

    Phylogenetic analysis confirmed that two genes required for acetoclastic methanogenesis, ackA and pta, were horizontally transferred to the ancestor of Methanosarcina from a derived cellulolytic organism in the class Clostridia. This event likely occurred within the last 475 million years, causing profound changes in planetary methane biogeochemistry. PMID:18055595

  16. Lateral gene transfer in a heavy metal-contaminated-groundwater microbial community

    DOE PAGES

    Hemme, Christopher L.; Green, Stefan J.; Rishishwar, Lavanya; ...

    2016-04-05

    Here, unraveling the drivers controlling the response and adaptation of biological communities to environmental change, especially anthropogenic activities, is a central but poorly understood issue in ecology and evolution. Comparative genomics studies suggest that lateral gene transfer (LGT) is a major force driving microbial genome evolution, but its role in the evolution of microbial communities remains elusive.

  17. Horizontal Gene Transfer among Bacteria and Its Role in Biological Evolution

    PubMed Central

    Arber, Werner

    2014-01-01

    This is a contribution to the history of scientific advance in the past 70 years concerning the identification of genetic information, its molecular structure, the identification of its functions and the molecular mechanisms of its evolution. Particular attention is thereby given to horizontal gene transfer among microorganisms, as well as to biosafety considerations with regard to beneficial applications of acquired scientific knowledge. PMID:25370194

  18. Bacteriophage-like Particles Associated with the Gene Transfer Agent of Methanococcus Voltale PS

    NASA Technical Reports Server (NTRS)

    Bertani, G.; Eiserling, F.; Pushkin, A.; Gingery, M.

    1999-01-01

    The methanogenic archaebacterium Methanococus voltae (strain PS) is known to produce a filterable, DNase resistant agent (called VTA, for voltae transfer agent), which carries very small fragments (4,400 base pairs) of bacterial DNA and is able to transduce bacterial genes between derivatives of the strain.

  19. Role of Vibrio cholerae exochitinase ChiA2 in horizontal gene transfer.

    PubMed

    Mondal, Moumita; Chatterjee, Nabendu Sekhar

    2016-03-01

    Vibrio cholerae exochitinase ChiA2 plays a key role in acquisition of nutrients by chitin hydrolysis in the natural environment as well as in pathogenesis in the intestinal milieu. In this study we demonstrate the importance of ChiA2 in horizontal gene transfer in the natural environment. We found that the expression of ChiA2 and TfoX, the central regulator of V. cholerae horizontal gene transfer, varied with changes in environmental conditions. The activity of ChiA2 was also dependent on these conditions. In 3 different environmental conditions tested here, we observed that the supporting environmental condition for maximum expression and activity of ChiA2 was 20 °C, pH 5.5, and 100 mmol/L salinity in the presence of chitin. The same condition also induced TfoX expression and was favorable for horizontal gene transfer in V. cholerae. High-performance liquid chromatography analysis showed that ChiA2 released a significant amount of (GlcNAc)2 from chitin hydrolysis under the favorable condition. We hypothesized that under the favorable environmental condition, ChiA2 was upregulated and maximally active to produce a significant amount of (GlcNAc)2 from chitin. The same environmental condition also induced tfoX expression, followed by its translational activation by the (GlcNAc)2 produced, leading to efficient horizontal gene transfer.

  20. Functional and Evolutionary Characterization of a Gene Transfer Agent’s Multilocus “Genome”

    PubMed Central

    Hynes, Alexander P.; Shakya, Migun; Mercer, Ryan G.; Grüll, Marc P.; Bown, Luke; Davidson, Fraser; Steffen, Ekaterina; Matchem, Heidi; Peach, Mandy E.; Berger, Tim; Grebe, Katherine; Zhaxybayeva, Olga; Lang, Andrew S.

    2016-01-01

    Gene transfer agents (GTAs) are phage-like particles that can package and transfer a random piece of the producing cell’s genome, but are unable to transfer all the genes required for their own production. As such, GTAs represent an evolutionary conundrum: are they selfish genetic elements propagating through an unknown mechanism, defective viruses, or viral structures “repurposed” by cells for gene exchange, as their name implies? In Rhodobacter capsulatus, production of the R. capsulatus GTA (RcGTA) particles is associated with a cluster of genes resembling a small prophage. Utilizing transcriptomic, genetic and biochemical approaches, we report that the RcGTA “genome” consists of at least 24 genes distributed across five distinct loci. We demonstrate that, of these additional loci, two are involved in cell recognition and binding and one in the production and maturation of RcGTA particles. The five RcGTA “genome” loci are widespread within Rhodobacterales, but not all loci have the same evolutionary histories. Specifically, two of the loci have been subject to frequent, probably virus-mediated, gene transfer events. We argue that it is unlikely that RcGTA is a selfish genetic element. Instead, our findings are compatible with the scenario that RcGTA is a virus-derived element maintained by the producing organism due to a selective advantage of within-population gene exchange. The modularity of the RcGTA “genome” is presumably a result of selection on the host organism to retain GTA functionality. PMID:27343288

  1. Cartilage degradation and invasion by rheumatoid synovial fibroblasts is inhibited by gene transfer of TIMP-1 and TIMP-3.

    PubMed

    van der Laan, W H; Quax, P H A; Seemayer, C A; Huisman, L G M; Pieterman, E J; Grimbergen, J M; Verheijen, J H; Breedveld, F C; Gay, R E; Gay, S; Huizinga, T W J; Pap, T

    2003-02-01

    Matrix metalloproteinases (MMPs) are believed to be pivotal enzymes in the invasion of articular cartilage by synovial tissue in rheumatoid arthritis (RA). Here, we investigated the effects of gene transfer of tissue inhibitors of metalloproteinases (TIMPs) on the invasiveness of RA synovial fibroblasts (RASF) in vitro and in vivo. Adenoviral vectors (Ad) were used for gene transfer. The effects of AdTIMP-1 and AdTIMP-3 gene transfer on matrix invasion were investigated in vitro in a transwell system. Cartilage invasion in vivo was studied in the SCID mouse co-implantation model for 60 days. In addition, the effects of AdTIMP-1 and AdTIMP-3 on cell proliferation were investigated. A significant reduction in invasiveness was demonstrated in vitro as well as in vivo in both the AdTIMP-1- and AdTIMP-3-transduced RASF compared with untransduced SF or SF that were transduced with control vectors. in vitro, the number of invading cells was reduced to 25% (P<0.001) in the AdTIMP-1-transduced cells and to 13% (P<0.0001) in the AdTIMP-3-transduced cells (% of untransduced cells). Cell proliferation was significantly inhibited by AdTIMP-3 and, less, by AdTIMP-1. In conclusion, overexpression of TIMP-1 and TIMP-3 by Ad gene transfer results in a marked reduction of the invasiveness of RASF in vitro and in the SCID mouse model. Apart from the inhibition of MMPs, a reduction in proliferation rate may contribute to this effect. These results suggest that overexpression of TIMPs, particularly TIMP-3 at the invasive front of pannus tissue, may provide a novel therapeutic strategy for inhibiting joint destruction in RA.

  2. Fat-to-glucose interconversion by hydrodynamic transfer of two glyoxylate cycle enzyme genes

    PubMed Central

    Cordero, P; Campion, J; Milagro, FI; Marzo, F; Martinez, JA

    2008-01-01

    The glyoxylate cycle, which is well characterized in higher plants and some microorganisms but not in vertebrates, is able to bypass the citric acid cycle to achieve fat-to-carbohydrate interconversion. In this context, the hydrodynamic transfer of two glyoxylate cycle enzymes, such as isocytrate lyase (ICL) and malate synthase (MS), could accomplish the shift of using fat for the synthesis of glucose. Therefore, 20 mice weighing 23.37 ± 0.96 g were hydrodinamically gene transferred by administering into the tail vein a bolus with ICL and MS. After 36 hours, body weight, plasma glucose, respiratory quotient and energy expenditure were measured. The respiratory quotient was increased by gene transfer, which suggests that a higher carbohydrate/lipid ratio is oxidized in such animals. This application could help, if adequate protocols are designed, to induce fat utilization for glucose synthesis, which might be eventually useful to reduce body fat depots in situations of obesity and diabetes. PMID:19077206

  3. Assessment of conjugal transfer of antibiotic resistance genes in Salmonella Typhimurium exposed to bile salts.

    PubMed

    He, Xinlong; Ahn, Juhee

    2014-08-01

    This study was designed to evaluate the transfer potential of antibiotic resistance genes in antibiotic-resistant Salmonella Typhimurium (S. Typhimurium(R)) in the presence of bile salts. The resistance of S. Typhimurium(R) to ampicillin, kanamycin, and tetracycline was increased by 64-, 64-, and 512-fold, respectively. The highest transfer frequency from S. Typhimurium(R) to Escherichia coli was observed at the bile salt concentration of 160 μg/ml (3.8 × 10(-3) transferrants/cells). The expression of traJ and traY was suppressed in S. Typhimurium(R) by bile salt. This study provides useful information for understanding the conjugative transfer of antibiotic resistance genes in S. Typhimurium under intestinal conditions.

  4. Radiative transfer in atmosphere-sea ice-ocean system

    SciTech Connect

    Jin, Z.; Stamnes, K.; Weeks, W.F.; Tsay, S.C.

    1996-04-01

    Radiative energy is critical in controlling the heat and mass balance of sea ice, which significantly affects the polar climate. In the polar oceans, light transmission through the atmosphere and sea ice is essential to the growth of plankton and algae and, consequently, to the microbial community both in the ice and in the ocean. Therefore, the study of radiative transfer in the polar atmosphere, sea ice, and ocean system is of particular importance. Lacking a properly coupled radiative transfer model for the atmosphere-sea ice-ocean system, a consistent study of the radiative transfer in the polar atmosphere, snow, sea ice, and ocean system has not been undertaken before. The radiative transfer processes in the atmosphere and in the ice and ocean have been treated separately. Because the radiation processes in the atmosphere, sea ice, and ocean depend on each other, this separate treatment is inconsistent. To study the radiative interaction between the atmosphere, clouds, snow, sea ice, and ocean, a radiative transfer model with consistent treatment of radiation in the coupled system is needed and is under development.

  5. Lunar transfer vehicle design issues with electric propulsion systems

    SciTech Connect

    Palaszewski, B.

    1989-01-01

    This paper describes parametric design studies of electric propulsion lunar transfer vehicles. In designing a lunar transfer vehicle, selecting the 'best' operating points for the design parameters allows significant reductions in the mass in low earth orbit (LEO) for the mission. These parameters include the specific impulse, the power level, and the propulsion technology. Many of the decisions regarding the operating points are controlled by the propulsion and power system technologies that are available for the spacecraft. The relationship between these technologies is discussed and analyzed here. It is found that both ion and MPD propulsion offer significant LEO mass reductions over O2/H2 for lunar transfer vehicle missions. The recommended operating points for the lunar transfer vehicle are an I(sp) of 5000 lb(f)-s/lb(m) and a 1 MW power level. For large lunar missions, krypton may be the best choice for ion propulsion. 17 refs.

  6. Definition of the ovalbumin gene promoter by transfer of an ovalglobin fusion gene into cultured cells.

    PubMed Central

    Knoll, B J; Zarucki-Schulz, T; Dean, D C; O'Malley, B W

    1983-01-01

    In order to study the initiation of transcription from the ovalbumin gene promoter, we constructed a hybrid gene (ovalglobin) in which 753 bps of ovalbumin gene 5'-flanking sequence were joined to the chicken adult beta-globin gene. When transfected into HeLa S3 cells, ovalglobin gene transcription initiated at the ovalbumin gene cap site, as measured by S1 nuclease and primer extension analysis. Deletion of 5'-flanking sequences to position -95 had little effect on transcription; deletion to -77 reduced transcription to about 20% of the wild type level and deletion to -48 reduced the level to about 2%. A deletion to -24, removing the sequence TATATAT, abolished transcription entirely. Hormonal regulation of the ovalglobin gene was observed when primary oviduct cells were used as recipients for DNA transfection. Under these conditions, addition of progesterone increased the level of ovalglobin transcripts to more than 10 times the uninduced level. Images PMID:6314256

  7. Targeted gene transfer to corneal endothelium in vivo by electric pulse.

    PubMed

    Oshima, Y; Sakamoto, T; Yamanaka, I; Nishi, T; Ishibashi, T; Inomata, H

    1998-10-01

    A novel method of in vivo targeted gene transfer to intentionally selected areas of the corneal endothelium was developed. Plasmid DNA with the lacZ gene coding for beta-galactosidase was injected into the anterior chamber of adult Wistar rats, and eight pulses of electricity at intensities ranging from 5 to 40 V/cm were delivered for 50 ms to the cornea with a specially designed electric probe in order to determine the effect of gene transfer on the corneal endothelial cells. Gene expression was visualized by enzymatic color reaction using X-gal in enucleated eyes on days 1, 3, 7, 14 and 21 after gene transfer. The treated eyes were then photographed and the X-gal-positive areas were evaluated by an image analyzer. The ratios of the areas (X-gal-positive area/area of entire corneal endothelium x 100%) were then calculated to determine gene transfection efficiency. The expression of beta-galactosidase was clearly detected in the cytoplasm of the corneal endothelial cells as early as day 1 and lasted until day 21. The most intense gene expression was observed on days 1 and 3 (5.21% on day 1 and 6.45% on day 3). The expression of beta-galactosidase on day 3 was most evident following delivery of 20 V electric pulses (0.09% at 5 V, 0.03% at 10 V, 6.45% at 20 V). beta-Galactosidase expression was limited to the corneal endothelial cells in highly selected areas and no beta-galactosidase expression was detected in any other intra- or axtraocular tissues. In addition, no cell damage was apparent in the cornea and no inflammation was detected in any other intraocular tissues. Thus, low-voltage electric pulses successfully transferred the gene of interest to highly selective areas of the corneal endothelium without inducing any pathological changes. This targeted gene transfer method appears to have great potential for use in gene therapy for ocular diseases.

  8. Evolution of genes and organisms: the tree/web of life in light of horizontal gene transfer.

    PubMed

    Olendzenski, Lorraine; Gogarten, J Peter

    2009-10-01

    Gene exchange necessitates expanding the model of the tree of life, impacts the notion of organismal and molecular most recent common ancestors, and provides examples of natural selection working at multiple levels. Gene exchange, whether by horizontal gene transfer (HGT), hybridization of species, or symbiosis, modifies the organismal tree of life into a web. Darwin suggested the tree of life was like a coral, where living surface branches were supported by masses of dead branches. In phylogenetic trees, organismal or molecular lineages coalesce back to a lucky universal ancestor whose descendents are found in current lineages and which coexisted with other, now-extinct lineages. HGT complicates the reconstruction of a universal ancestor; genes in a genome can have different evolutionary histories, and even infrequent gene transfer will cause different molecular lineages to coalesce to molecular ancestors that existed in different organismal lineages and at different times. HGT, as well as symbiosis, provides a mechanism for integrating and expanding the organizational level on which natural selection acts, contributing to selection at the group and community level.

  9. Method of measuring heat influx of a cryogenic transfer system

    DOEpatents

    Niemann, Ralph C.; Zelipsky, Steven A.; Rezmer, Ronald R.; Smelser, Peter

    1981-01-01

    A method is provided for measuring the heat influx of a cryogenic transfer system. A gaseous phase of the cryogen used during normal operation of the system is passed through the system. The gaseous cryogen at the inlet to the system is tempered to duplicate the normal operating temperature of the system inlet. The temperature and mass flow rate of the gaseous cryogen is measured at the outlet of the system, and the heat capacity of the cryogen is determined. The heat influx of the system is then determined from known thermodynamic relationships.

  10. Advanced two-phase heat transfer systems

    NASA Technical Reports Server (NTRS)

    Swanson, Theodore D.

    1992-01-01

    Future large spacecraft, such as the Earth Observing System (EOS) platforms, will require a significantly more capable thermal control system than is possible with current 'passive' technology. Temperatures must be controlled much more tightly over a larger surface area. Numerous heat load sources will often be located inside the body of the spacecraft without a good view to space. Power levels and flux densities may be higher than can be accommodated with traditional technology. Integration and ground testing will almost certainly be much more difficult with such larger, more complex spacecraft. For these and similar reasons, the Goddard Space Flight Center (GSFC) has been developing a new, more capable thermal control technology called capillary pumped loops (CPL's). CPL's represent an evolutionary improvement over heat pipes; they can transport much greater quantities of heat over much longer distances and can serve numerous heat load sources. In addition, CPL's can be fabricated into large cold plates that can be held to tight thermal gradients. Development of this technology began in the early 1980's and is now reaching maturity. CPL's have recently been baselined for the EOS-AM platform (1997 launch) and the COMET spacecraft (1992 launch). This presentation describes this new technology and its applications. Most of the viewgraphs are self descriptive. For those that are less clear additional comments are provided.

  11. The case of horizontal gene transfer from bacteria to the peculiar dinoflagellate plastid genome

    PubMed Central

    Mackiewicz, Paweł; Bodył, Andrzej; Moszczyński, Krzysztof

    2013-01-01

    Organelle genomes lose their genes by transfer to host nuclear genomes, but only occasionally are enriched by foreign genes from other sources. In contrast to mitochondria, plastid genomes are especially resistant to such horizontal gene transfer (HGT), and thus every gene acquired in this way is notable. An exceptional case of HGT was recently recognized in the peculiar peridinin plastid genome of dinoflagellates, which is organized in plasmid-like minicircles. Genomic and phylogenetic analyses of Ceratium horridum and Pyrocystis lunula minicircles revealed four genes and one unannotated open reading frame that probably were gained from bacteria belonging to the Bacteroidetes. Such bacteria seem to be a good source of genes because close endosymbiotic associations between them and dinoflagellates have been observed. The HGT-acquired genes are involved in plastid functions characteristic of other photosynthetic eukaryotes, and their arrangement resembles bacterial operons. These studies indicate that the peridinin plastid genome, usually regarded as having resulted from reduction and fragmentation of a typical plastid genome derived from red algae, may have a chimeric origin that includes bacterial contributions. Potential contamination of the Ceratium and Pyrocystis plastid genomes by bacterial sequences and the controversial localization of their minicircles in the nucleus are also discussed. PMID:24195014

  12. Gene transfer in the liver using recombinant adeno-associated virus

    PubMed Central

    Ahmed, Seemin Seher; Li, Jia; Godwin, Jonathan; Gao, Guangping; Zhong, Li

    2013-01-01

    Liver-directed gene transfer and gene therapy are rapidly gaining attention primarily because the liver is centrally involved in a variety of metabolic functions that are affected in various inherited disorders. Recombinant adeno-associated virus (rAAV) is a popular gene