WorldWideScience

Sample records for genetically engineered mesenchymal

  1. Genetic Engineering of Mesenchymal Stem Cells for Regenerative Medicine.

    Science.gov (United States)

    Nowakowski, Adam; Walczak, Piotr; Janowski, Miroslaw; Lukomska, Barbara

    2015-10-01

    Mesenchymal stem cells (MSCs), which can be obtained from various organs and easily propagated in vitro, are one of the most extensively used types of stem cells and have been shown to be efficacious in a broad set of diseases. The unique and highly desirable properties of MSCs include high migratory capacities toward injured areas, immunomodulatory features, and the natural ability to differentiate into connective tissue phenotypes. These phenotypes include bone and cartilage, and these properties predispose MSCs to be therapeutically useful. In addition, MSCs elicit their therapeutic effects by paracrine actions, in which the metabolism of target tissues is modulated. Genetic engineering methods can greatly amplify these properties and broaden the therapeutic capabilities of MSCs, including transdifferentiation toward diverse cell lineages. However, cell engineering can also affect safety and increase the cost of therapy based on MSCs; thus, the advantages and disadvantages of these procedures should be discussed. In this review, the latest applications of genetic engineering methods for MSCs with regenerative medicine purposes are presented.

  2. Genetic engineering of mesenchymal stem cells and its application in human disease therapy.

    Science.gov (United States)

    Hodgkinson, Conrad P; Gomez, José A; Mirotsou, Maria; Dzau, Victor J

    2010-11-01

    The use of stem cells for tissue regeneration and repair is advancing both at the bench and bedside. Stem cells isolated from bone marrow are currently being tested for their therapeutic potential in a variety of clinical conditions including cardiovascular injury, kidney failure, cancer, and neurological and bone disorders. Despite the advantages, stem cell therapy is still limited by low survival, engraftment, and homing to damage area as well as inefficiencies in differentiating into fully functional tissues. Genetic engineering of mesenchymal stem cells is being explored as a means to circumvent some of these problems. This review presents the current understanding of the use of genetically engineered mesenchymal stem cells in human disease therapy with emphasis on genetic modifications aimed to improve survival, homing, angiogenesis, and heart function after myocardial infarction. Advancements in other disease areas are also discussed.

  3. Genetic Engineering of Mesenchymal Stem Cells and Its Application in Human Disease Therapy

    OpenAIRE

    Hodgkinson, Conrad P; Gomez, José A.; Mirotsou, Maria; Dzau, Victor J.

    2010-01-01

    Hodgkinson and colleagues review the current status of knowledge with respect to the genetic modifications being explored as a means to improve mesenchymal stem cell therapy for human diseases, with a particular focus on cardiovascular diseases.

  4. Lentiviral Mediating Genetic Engineered Mesenchymal Stem Cells for Releasing IL-27 as a Gene Therapy Approach for Autoimmune Diseases

    OpenAIRE

    Shohreh Hajizadeh-Sikaroodi; Ahmad Hosseini; Ali Falla; Hajar Estiri; Zahra Noormohammadi; Mohammad Salehi; Sayyed Mohammad Hossein Ghaderian; Haleh Akhavan Niaki; Masoud Soleimani; Bahram Kazemi

    2014-01-01

    Objective: Autoimmune diseases precede a complex dysregulation of the immune system. T helper17 (Th17) and interleukin (IL)-17 have central roles in initiation of inflammation and subsequent autoimmune diseases. IL-27 significantly controls autoimmune diseases by Th17 and IL-17 suppression. In the present study we have created genetic engineered mesenchymal stem cells (MSCs) that mediate with lentiviral vectors to release IL-27 as an adequate vehicle for ex vivo gene therapy in...

  5. Genetic engineering of mesenchymal stromal cells for cancer therapy: turning partners in crime into Trojan horses

    Directory of Open Access Journals (Sweden)

    Niess Hanno

    2016-09-01

    Full Text Available Mesenchymal stromal cells (MSCs are adult progenitor cells with a high migratory and differentiation potential, which influence a broad range of biological functions in almost every tissue of the body. Among other mechanisms, MSCs do so by the secretion of molecular cues, differentiation toward more specialized cell types, or influence on the immune system. Expanding tumors also depend on the contribution of MSCs to building a supporting stroma, but the effects of MSCs appear to go beyond the mere supply of connective tissues. MSCs show targeted “homing” toward growing tumors, which is then followed by exerting direct and indirect effects on cancer cells. Several research groups have developed novel strategies that make use of the tumor tropism of MSCs by engineering them to express a transgene that enables an attack on cancer growth. This review aims to familiarize the reader with the current knowledge about MSC biology, the existing evidence for MSC contribution to tumor growth with its underlying mechanisms, and the strategies that have been developed using MSCs to deploy an anticancer therapy.

  6. Genetically engineered theranostic mesenchymal stem cells for the evaluation of the anticancer efficacy of enzyme/prodrug systems.

    Science.gov (United States)

    Nouri, Faranak Salman; Wang, Xing; Hatefi, Arash

    2015-02-28

    Over the past decade, various enzyme/prodrug systems such as thymidine kinase/ganciclovir (TK/GCV), yeast cytosine deaminase/5-fluorocytosine (yCD/5-FC) and nitroreductase/CB1954 (NTR/CB1954) have been used for stem cell mediated suicide gene therapy of cancer. Yet, no study has been conducted to compare and demonstrate the advantages and disadvantages of using one system over another. Knowing that each enzyme/prodrug system has its own strengths and weaknesses, we utilized mesenchymal stem cells (MSCs) as a medium to perform for the first time a comparative study that illustrated the impact of subtle differences among these systems on the therapeutic outcome. For therapeutic purposes, we first genetically modified MSCs to stably express a panel of four suicide genes including TK (TK007 and TK(SR39) mutants), yeast cytosine deaminase:uracil phosphoribosyltransferase (yCD:UPRT) and nitroreductase (NTR). Then, we evaluated the anticancer efficacies of the genetically engineered MSCs in vitro and in vivo by using SKOV3 cell line which is sensitive to all four enzyme/prodrug systems. In addition, all MSCs were engineered to stably express luciferase gene making them suitable for quantitative imaging and dose-response relationship studies in animals. Considering the limitations imposed by the prodrugs' bystander effects, our findings show that yCD:UPRT/5-FC is the most effective enzyme/prodrug system among the ones tested. Our findings also demonstrate that theranostic MSCs are a reliable medium for the side-by-side evaluation and screening of the enzyme/prodrug systems at the preclinical level. The results of this study could help scientists who utilize cell-based, non-viral or viral vectors for suicide gene therapy of cancer make more informed decisions when choosing enzyme/prodrug systems.

  7. Clinical trial perspective for adult and juvenile Huntington′s disease using genetically-engineered mesenchymal stem cells

    Directory of Open Access Journals (Sweden)

    Peter Deng

    2016-01-01

    Full Text Available Progress to date from our group and others indicate that using genetically-engineered mesenchymal stem cells (MSC to secrete brain-derived neurotrophic factor (BDNF supports our plan to submit an Investigational New Drug application to the Food and Drug Administration for the future planned Phase 1 safety and tolerability trial of MSC/BDNF in patients with Huntington′s disease (HD. There are also potential applications of this approach beyond HD. Our biological delivery system for BDNF sets the precedent for adult stem cell therapy in the brain and could potentially be modified for other neurodegenerative disorders such as amyotrophic lateral sclerosis (ALS, spinocerebellar ataxia (SCA, Alzheimer′s disease, and some forms of Parkinson′s disease. The MSC/BDNF product could also be considered for studies of regeneration in traumatic brain injury, spinal cord and peripheral nerve injury. This work also provides a platform for our future gene editing studies, since we will again use MSCs to deliver the needed molecules into the central nervous system.

  8. Clinical trial perspective for adult and juvenile Huntington’s disease using genetically-engineered mesenchymal stem cells

    Institute of Scientific and Technical Information of China (English)

    Peter Deng; Audrey Torrest; Kari Pollock; Heather Dahlenburg; Geralyn Annett; Jan A. Nolta; Kyle D. Fink

    2016-01-01

    Progress to date from our group and others indicate that using genetically-engineered mesenchymal stem cells (MSC) to secrete brain-derived neurotrophic factor (BDNF) supports our plan to submit an Investigational New Drug application to the Food and Drug Administration for the future planned Phase 1 safety and tolera-bility trial of MSC/BDNF in patients with Huntington’s disease (HD). There are also potential applications of this approach beyond HD. Our biological delivery system for BDNF sets the precedent for adult stem cell ther-apy in the brain and could potentially be modiifed for other neurodegenerative disorders such as amyotrophic lateral sclerosis (ALS), spinocerebellar ataxia (SCA), Alzheimer’s disease, and some forms of Parkinson’s dis-ease. The MSC/BDNF product could also be considered for studies of regeneration in traumatic brain injury, spinal cord and peripheral nerve injury. This work also provides a platform for our future gene editing studies, since we will again use MSCs to deliver the needed molecules into the central nervous system.

  9. Genetic Engineering

    Science.gov (United States)

    Phillips, John

    1973-01-01

    Presents a review of genetic engineering, in which the genotypes of plants and animals (including human genotypes) may be manipulated for the benefit of the human species. Discusses associated problems and solutions and provides an extensive bibliography of literature relating to genetic engineering. (JR)

  10. Lentiviral Mediating Genetic Engineered Mesenchymal Stem Cells for Releasing IL-27 as a Gene Therapy Approach for Autoimmune Diseases

    Directory of Open Access Journals (Sweden)

    Shohreh Hajizadeh-Sikaroodi

    2014-04-01

    Full Text Available Objective: Autoimmune diseases precede a complex dysregulation of the immune system. T helper17 (Th17 and interleukin (IL-17 have central roles in initiation of inflammation and subsequent autoimmune diseases. IL-27 significantly controls autoimmune diseases by Th17 and IL-17 suppression. In the present study we have created genetic engineered mesenchymal stem cells (MSCs that mediate with lentiviral vectors to release IL-27 as an adequate vehicle for ex vivo gene therapy in the reduction of inflammation and autoimmune diseases. Materials and Methods: In this experimental study, we isolated adipose-derived MSCs (AD-MSCs from lipoaspirate and subsequently characterized them by differentiation. Two subunits of IL-27 (p28 and EBI3 were cloned in a pCDH-513B-1 lentiviral vector. Expressions of p28 and EBI3 (Epstein-Barr virus induced gene 3 were determined by real time polymerase chain reaction (PCR. MSCs were transduced by a pCDH-CMV-p28-IRESEBI3- EF-copGFP-Pur lentiviral vector and the bioassay of IL-27 was evaluated by IL-10 expression. Results: Cell differentiation confirmed true isolation of MSCs from lipoaspirate. Restriction enzyme digestion and sequencing verified successful cloning of both p28 and EBI3 in the pCDH-513B-1 lentiviral vector. Real time PCR showed high expressions level of IL-27 and IL-10 as well as accurate activity of IL-27. Conclusion: The results showed transduction of functional IL-27 to AD-MSCs by means of a lentiviral vector. The lentiviral vector did not impact MSC characteristics.

  11. Fluorescence molecular tomography enables in vivo visualization and quantification of nonunion fracture repair induced by genetically engineered mesenchymal stem cells.

    Science.gov (United States)

    Zilberman, Yoram; Kallai, Ilan; Gafni, Yossi; Pelled, Gadi; Kossodo, Sylvie; Yared, Wael; Gazit, Dan

    2008-04-01

    Fluorescence molecular tomography (FMT) is a novel tomographic near-infrared (NIR) imaging modality that enables 3D quantitative determination of fluorochrome distribution in tissues of live small animals at any depth. This study demonstrates a noninvasive, quantitative method of monitoring engineered bone remodeling via FMT. Murine mesenchymal stem cells overexpressing the osteogenic gene BMP2 (mMSCs-BMP2) were implanted into the thigh muscle and into a radial nonunion bone defect model in C3H/HeN mice. Real-time imaging of bone formation was performed following systemic administration of the fluorescent bisphosphonate imaging agent OsteoSense, an hydroxyapatite-directed bone-imaging probe. The mice underwent imaging on days 7, 14, and 21 postimplantation. New bone formation at the implantation sites was quantified using micro-computed tomography (micro-CT) imaging. A higher fluorescent signal occurred at the site of the mMSC-BMP2 implants than that found in controls. Micro-CT imaging revealed a mass of mature bone formed in the implantation sites on day 21, a finding also confirmed by histology. These findings highlight the effectiveness of FMT as a functional platform for molecular imaging in the field of bone regeneration and tissue engineering.

  12. The effect of lentivirus-mediated PSPN genetic engineering bone marrow mesenchymal stem cells on Parkinson's disease rat model

    National Research Council Canada - National Science Library

    Yin, Xiaofeng; Xu, Huamin; Jiang, Yunxia; Deng, Wenshuai; Wu, Zeyu; Xiang, Hengwei; Sun, Peng; Xie, Junxia

    2014-01-01

    ...) which have been found to promote the survival of specific populations of neurons. The aim of this study was to assess the potential therapeutic function of gene-modified mesenchymal stem cells (MSCs...

  13. Mesenchymal Stem Cells and Tooth Engineering

    Institute of Scientific and Technical Information of China (English)

    Li Peng; Ling Ye; Xue-dong Zhou

    2009-01-01

    Tooth loss compromises human oral health. Although several prosthetic methods, such as artificial denture and dental implants, are clinical therapies to tooth loss problems, they are thought to have safety and usage time issues. Recently, tooth tissue engineering has attracted more and more attention. Stem cell based tissue engineering is thought to be a promising way to replace the missing tooth. Mesenchymal stem cells (MSCs) are multipotent stem cells which can differentiate into a variety of cell types. The potential MSCs for tooth regeneration mainly include stem cells from human exfoliated deciduous teeth (SHEDs), adult dental pulp stem cells (DPSCs), stem cells from the apical part of the papilla (SCAPs), stem cells from the dental follicle (DFSCs), periodontal ligament stem cells (PDLSCs) and bone marrow derived mesenchymal stem cells (BMSCs). This review outlines the recent progress in the mesenchymal stem cells used in tooth regeneration.

  14. Cartilage Engineering from Mesenchymal Stem Cells

    Science.gov (United States)

    Goepfert, C.; Slobodianski, A.; Schilling, A. F.; Adamietz, P.; Pörtner, R.

    Mesenchymal progenitor cells known as multipotent mesenchymal stromal cells or mesenchymal stem cells (MSC) have been isolated from various tissues. Since they are able to differentiate along the mesenchymal lineages of cartilage and bone, they are regarded as promising sources for the treatment of skeletal defects. Tissue regeneration in the adult organism and in vitro engineering of tissues is hypothesized to follow the principles of embryogenesis. The embryonic development of the skeleton has been studied extensively with respect to the regulatory mechanisms governing morphogenesis, differentiation, and tissue formation. Various concepts have been designed for engineering tissues in vitro based on these developmental principles, most of them involving regulatory molecules such as growth factors or cytokines known to be the key regulators in developmental processes. Growth factors most commonly used for in vitro cultivation of cartilage tissue belong to the fibroblast growth factor (FGF) family, the transforming growth factor-beta (TGF-β) super-family, and the insulin-like growth factor (IGF) family. In this chapter, in vivo actions of members of these growth factors described in the literature are compared with in vitro concepts of cartilage engineering making use of these growth factors.

  15. Genetically engineered foods

    Science.gov (United States)

    Bioengineered foods; GMOs; Genetically modified foods ... helps speed up the process of creating new foods with desired traits. The possible benefits of genetic engineering include: More nutritious food Tastier food Disease- and ...

  16. Genetically Engineered Cyanobacteria

    Science.gov (United States)

    Zhou, Ruanbao (Inventor); Gibbons, William (Inventor)

    2015-01-01

    The disclosed embodiments provide cyanobacteria spp. that have been genetically engineered to have increased production of carbon-based products of interest. These genetically engineered hosts efficiently convert carbon dioxide and light into carbon-based products of interest such as long chained hydrocarbons. Several constructs containing polynucleotides encoding enzymes active in the metabolic pathways of cyanobacteria are disclosed. In many instances, the cyanobacteria strains have been further genetically modified to optimize production of the carbon-based products of interest. The optimization includes both up-regulation and down-regulation of particular genes.

  17. Engineering mesenchymal stem cells for regenerative medicine and drug delivery.

    Science.gov (United States)

    Park, Ji Sun; Suryaprakash, Smruthi; Lao, Yeh-Hsing; Leong, Kam W

    2015-08-01

    Researchers have applied mesenchymal stem cells (MSC) to a variety of therapeutic scenarios by harnessing their multipotent, regenerative, and immunosuppressive properties with tropisms toward inflamed, hypoxic, and cancerous sites. Although MSC-based therapies have been shown to be safe and effective to a certain degree, the efficacy remains low in most cases when MSC are applied alone. To enhance their therapeutic efficacy, researchers have equipped MSC with targeted delivery functions using genetic engineering, therapeutic agent incorporation, and cell surface modification. MSC can be genetically modified virally or non-virally to overexpress therapeutic proteins that complement their innate properties. MSC can also be primed with non-peptidic drugs or magnetic nanoparticles for enhanced efficacy and externally regulated targeting, respectively. Furthermore, MSC can be functionalized with targeting moieties to augment their homing toward therapeutic sites using enzymatic modification, chemical conjugation, or non-covalent interactions. These engineering techniques are still works in progress, requiring optimization to improve the therapeutic efficacy and targeting effectiveness while minimizing any loss of MSC function. In this review, we will highlight the advanced techniques of engineering MSC, describe their promise and the challenges of translation into clinical settings, and suggest future perspectives on realizing their full potential for MSC-based therapy. Copyright © 2015 Elsevier Inc. All rights reserved.

  18. Genetically Engineering Entomopathogenic Fungi.

    Science.gov (United States)

    Zhao, H; Lovett, B; Fang, W

    2016-01-01

    Entomopathogenic fungi have been developed as environmentally friendly alternatives to chemical insecticides in biocontrol programs for agricultural pests and vectors of disease. However, mycoinsecticides currently have a small market share due to low virulence and inconsistencies in their performance. Genetic engineering has made it possible to significantly improve the virulence of fungi and their tolerance to adverse conditions. Virulence enhancement has been achieved by engineering fungi to express insect proteins and insecticidal proteins/peptides from insect predators and other insect pathogens, or by overexpressing the pathogen's own genes. Importantly, protein engineering can be used to mix and match functional domains from diverse genes sourced from entomopathogenic fungi and other organisms, producing insecticidal proteins with novel characteristics. Fungal tolerance to abiotic stresses, especially UV radiation, has been greatly improved by introducing into entomopathogens a photoreactivation system from an archaean and pigment synthesis pathways from nonentomopathogenic fungi. Conversely, gene knockout strategies have produced strains with reduced ecological fitness as recipients for genetic engineering to improve virulence; the resulting strains are hypervirulent, but will not persist in the environment. Coupled with their natural insect specificity, safety concerns can also be mitigated by using safe effector proteins with selection marker genes removed after transformation. With the increasing public concern over the continued use of synthetic chemical insecticides and growing public acceptance of genetically modified organisms, new types of biological insecticides produced by genetic engineering offer a range of environmentally friendly options for cost-effective control of insect pests. Copyright © 2016 Elsevier Inc. All rights reserved.

  19. Genetic and epigenetic effect of estrogen on mesenchymal stem cell maintenance and differentiation

    OpenAIRE

    2013-01-01

    Ankara : The Department of Molecular Biology and Genetics and the Graduate School of Engineering and Science of Bilkent Univ., 2013. Thesis (Ph. D.) -- Bilkent University, 2013. Includes bibliographical references leaves 126-143. Mesenchymal stem cells (MSCs) have the potential to differentiate into multiple cell types and immune privileged characteristics. These features make MSCs a hope in tissue engineering and cell based treatment applications. Tremendous amount of stud...

  20. Paper Genetic Engineering.

    Science.gov (United States)

    MacClintic, Scott D.; Nelson, Genevieve M.

    Bacterial transformation is a commonly used technique in genetic engineering that involves transferring a gene of interest into a bacterial host so that the bacteria can be used to produce large quantities of the gene product. Although several kits are available for performing bacterial transformation in the classroom, students do not always…

  1. Selected Readings in Genetic Engineering

    Science.gov (United States)

    Mertens, Thomas R.; Robinson, Sandra K.

    1973-01-01

    Describes different sources of readings for understanding issues and concepts of genetic engineering. Broad categories of reading materials are: concerns about genetic engineering; its background; procedures; and social, ethical and legal issues. References are listed. (PS)

  2. Safe genetically engineered plants

    Science.gov (United States)

    Rosellini, D.; Veronesi, F.

    2007-10-01

    The application of genetic engineering to plants has provided genetically modified plants (GMPs, or transgenic plants) that are cultivated worldwide on increasing areas. The most widespread GMPs are herbicide-resistant soybean and canola and insect-resistant corn and cotton. New GMPs that produce vaccines, pharmaceutical or industrial proteins, and fortified food are approaching the market. The techniques employed to introduce foreign genes into plants allow a quite good degree of predictability of the results, and their genome is minimally modified. However, some aspects of GMPs have raised concern: (a) control of the insertion site of the introduced DNA sequences into the plant genome and of its mutagenic effect; (b) presence of selectable marker genes conferring resistance to an antibiotic or an herbicide, linked to the useful gene; (c) insertion of undesired bacterial plasmid sequences; and (d) gene flow from transgenic plants to non-transgenic crops or wild plants. In response to public concerns, genetic engineering techniques are continuously being improved. Techniques to direct foreign gene integration into chosen genomic sites, to avoid the use of selectable genes or to remove them from the cultivated plants, to reduce the transfer of undesired bacterial sequences, and make use of alternative, safer selectable genes, are all fields of active research. In our laboratory, some of these new techniques are applied to alfalfa, an important forage plant. These emerging methods for plant genetic engineering are briefly reviewed in this work.

  3. Genetic engineering of cyanobacteria

    DEFF Research Database (Denmark)

    Jacobsen, Jacob Hedemand

    , including genetic tools that allow metabolic engineering. The cyanobacterial phylum represents a diverse group of aerobic photosynthetic bacteria that are widespread in nature. Cyanobacteria shaped our atmosphere by oxygen evolution through the splitting of water using energy from sunlight. The sole carbon...... and characterized for growth phenotype and glycogen content. While no difference in growth rate or glycogen content was detected between the phosphorylase double mutant and wild type strain, we found that both glycogen phophyrylases must be genetically inactivated to eliminate glycogen phosphorylase activity...

  4. Genetic engineering in biotechnology

    Energy Technology Data Exchange (ETDEWEB)

    Bedate, C.A.; Morales, J.C.; Lopez, E.H.

    1981-09-01

    The objective of this book is to encourage the use of genetic engineering for economic development. The report covers: (1) Precedents of genetic engineering; (2) a brief description of the technology, including the transfer of DNA in bacteria (vectors, E. coli and B. subtilis hosts, stages, and technical problems), practical examples of techniques used and their products (interferon; growth hormone; insulin; treatment of blood cells, Talasemia, and Lesch-Nyhan syndrome; and more nutritious soya), transfer to higher organisms, and cellular fusion; (3) biological risks and precautions; (4) possible applications (production of hydrogen, hydrocarbons, alcohol, chemicals, enzymes, peptides, viral antigens, monoclonal antibodies, genes, proteins, and insecticides; metal extraction; nitrogen fixation; biodegradation; and new varieties of plants and animals; and (5) international activities.

  5. Genetic engineering of cyanobacteria

    DEFF Research Database (Denmark)

    Jacobsen, Jacob Hedemand

    , including genetic tools that allow metabolic engineering. The cyanobacterial phylum represents a diverse group of aerobic photosynthetic bacteria that are widespread in nature. Cyanobacteria shaped our atmosphere by oxygen evolution through the splitting of water using energy from sunlight. The sole carbon...... and its natural ability to take up and stably integrate heterologous DNA make Synechococcus sp. PCC 7002 a good candidate for metabolic engineering. For targeted gene inactivation, a suite of vectors were made by adaptation of a system previously used in plants and fungi. The vectors include a cassette...... and characterized for growth phenotype and glycogen content. While no difference in growth rate or glycogen content was detected between the phosphorylase double mutant and wild type strain, we found that both glycogen phophyrylases must be genetically inactivated to eliminate glycogen phosphorylase activity...

  6. Cell surface engineering of mesenchymal stem cells.

    Science.gov (United States)

    Sarkar, Debanjan; Zhao, Weian; Gupta, Ashish; Loh, Wei Li; Karnik, Rohit; Karp, Jeffrey M

    2011-01-01

    By leveraging the capacity to promote regeneration, stem cell therapies offer enormous hope for solving some of the most tragic illnesses, diseases, and tissue defects world-wide. However, a significant barrier to the effective implementation of cell therapies is the inability to target a large quantity of viable cells with high efficiency to tissues of interest. Systemic infusion is desired as it minimizes the invasiveness of cell therapy, and maximizes practical aspects of repeated doses. However, cell types such as mesenchymal stem cells exhibit a poor homing capability or lose their capacity to home following culture expansion (i.e. FASEB J 21:3197-3207, 2007; Circulation 108:863-868, 2003; Stroke: A Journal of Cerebral Circulation 32:1005-1011; Blood 104:3581-3587, 2004). To address this challenge, we have developed a simple platform technology to chemically attach cell adhesion molecules to the cell surface to improve the homing efficiency to specific tissues. This chemical approach involves a stepwise process including (1) treatment of cells with sulfonated biotinyl-N-hydroxy-succinimide to introduce biotin groups on the cell surface, (2) addition of streptavidin that binds to the biotin on the cell surface and presents unoccupied binding sites, and (3) attachment of biotinylated targeting ligands that promote adhesive interactions with vascular endothelium. Specifically, in our model system, a biotinylated cell rolling ligand, sialyl Lewisx (SLeX), found on the surface of leukocytes (i.e., the active site of the P-selectin glycoprotein ligand (PSGL-1)), is conjugated on MSC surface. The SLeX engineered MSCs exhibit a rolling response on a P-selectin coated substrate under shear stress conditions. This indicates that this approach can be used to potentially target P-selectin expressing endothelium in the more marrow or at sites of inflammation. Importantly, the surface modification has no adverse impact on MSCs' native phenotype including their multilineage

  7. Training human mesenchymal stromal cells for bone tissue engineering applications

    NARCIS (Netherlands)

    Doorn, J.

    2012-01-01

    Human mesenchymal stromal cells (hMSCs) are an interesting source for cell therapies and tissue engineering applications, because these cells are able to differentiate into various target tissues, such as bone, cartilage, fat and endothelial cells. In addition, they secrete a wide array of growth fa

  8. A genetic engineering approach to genetic algorithms.

    Science.gov (United States)

    Gero, J S; Kazakov, V

    2001-01-01

    We present an extension to the standard genetic algorithm (GA), which is based on concepts of genetic engineering. The motivation is to discover useful and harmful genetic materials and then execute an evolutionary process in such a way that the population becomes increasingly composed of useful genetic material and increasingly free of the harmful genetic material. Compared to the standard GA, it provides some computational advantages as well as a tool for automatic generation of hierarchical genetic representations specifically tailored to suit certain classes of problems.

  9. Moral Fantasy in Genetic Engineering.

    Science.gov (United States)

    Boone, C. Keith

    1984-01-01

    Discusses the main ethical issues generated by the new genetics and suggests ways to think about them. Concerns include "playing God," violation of the natural order of the universe, and abuse of genetic technology. Critical distinctions for making difficult decisions about genetic engineering issues are noted. (DH)

  10. Genetic engineering, medicine and medical genetics.

    Science.gov (United States)

    Motulsky, A G

    1984-01-01

    The impact of DNA technology in the near future will be on the manufacture of biologic agents and reagents that will lead to improved therapy and diagnosis. The use of DNA technology for prenatal and preclinical diagnosis in genetic diseases is likely to affect management of genetic diseases considerably. New and old questions regarding selective abortion and the psychosocial impact of early diagnosis of late appearing diseases and of genetic susceptibilities are being raised. Somatic therapy with isolated genes to treat disease has not been achieved. True germinal genetic engineering is far off for humans but may find applications in animal agriculture.

  11. Genetic and metabolic engineering

    OpenAIRE

    Yang,Yea-Tyng; Bennett, George N.; San, Ka-yiu

    1998-01-01

    Recent advances in molecular biology techniques, analytical methods and mathematical tools have led to a growing interest in using metabolic engineering to redirect metabolic fluxes for industrial and medical purposes. Metabolic engineering is referred to as the directed improvement of cellular properties through the modification of specific biochemical reactions or the introduction of new ones, with the use of recombinant DNA technology (Stephanopoulos, 1999). This multidisciplinary field dr...

  12. Genetically engineered yeast

    DEFF Research Database (Denmark)

    2014-01-01

    A genetically modified Saccharomyces cerevisiae comprising an active fermentation pathway producing 3-HP expresses an exogenous gene expressing the aminotransferase YhxA from Bacillus cereus AH1272 catalysing a transamination reaction between beta-alanine and pyruvate to produce malonate semialde......A genetically modified Saccharomyces cerevisiae comprising an active fermentation pathway producing 3-HP expresses an exogenous gene expressing the aminotransferase YhxA from Bacillus cereus AH1272 catalysing a transamination reaction between beta-alanine and pyruvate to produce malonate...

  13. Genetic Engineering Workshop Report, 2010

    Energy Technology Data Exchange (ETDEWEB)

    Allen, J; Slezak, T

    2010-11-03

    The Lawrence Livermore National Laboratory (LLNL) Bioinformatics group has recently taken on a role in DTRA's Transformation Medical Technologies (TMT) program. The high-level goal of TMT is to accelerate the development of broad-spectrum countermeasures. To achieve this goal, there is a need to assess the genetic engineering (GE) approaches, potential application as well as detection and mitigation strategies. LLNL was tasked to coordinate a workshop to determine the scope of investments that DTRA should make to stay current with the rapid advances in genetic engineering technologies, so that accidental or malicious uses of GE technologies could be adequately detected and characterized. Attachment A is an earlier report produced by LLNL for TMT that provides some relevant background on Genetic Engineering detection. A workshop was held on September 23-24, 2010 in Springfield, Virginia. It was attended by a total of 55 people (see Attachment B). Twenty four (44%) of the attendees were academic researchers involved in GE or bioinformatics technology, 6 (11%) were from DTRA or the TMT program management, 7 (13%) were current TMT performers (including Jonathan Allen and Tom Slezak of LLNL who hosted the workshop), 11 (20%) were from other Federal agencies, and 7 (13%) were from industries that are involved in genetic engineering. Several attendees could be placed in multiple categories. There were 26 attendees (47%) who were from out of the DC area and received travel assistance through Invitational Travel Orders (ITOs). We note that this workshop could not have been as successful without the ability to invite experts from outside of the Beltway region. This workshop was an unclassified discussion of the science behind current genetic engineering capabilities. US citizenship was not required for attendance. While this may have limited some discussions concerning risk, we felt that it was more important for this first workshop to focus on the scientific state of

  14. Genetic engineering of Geobacillus spp.

    Science.gov (United States)

    Kananavičiūtė, Rūta; Čitavičius, Donaldas

    2015-04-01

    Members of the genus Geobacillus are thermophiles that are of great biotechnological importance, since they are sources of many thermostable enzymes. Because of their metabolic versatility, geobacilli can be used as whole-cell catalysts in processes such as bioconversion and bioremediation. The effective employment of Geobacillus spp. requires the development of reliable methods for genetic engineering of these bacteria. Currently, genetic manipulation tools and protocols are under rapid development. However, there are several convenient cloning vectors, some of which replicate autonomously, while others are suitable for the genetic modification of chromosomal genes. Gene expression systems are also intensively studied. Combining these tools together with proper techniques for DNA transfer, some Geobacillus strains were shown to be valuable producers of recombinant proteins and industrially important biochemicals, such as ethanol or isobutanol. This review encompasses the progress made in the genetic engineering of Geobacillus spp. and surveys the vectors and transformation methods that are available for this genus.

  15. "Genetically Engineered" Nanoelectronics

    Science.gov (United States)

    Klimeck, Gerhard; Salazar-Lazaro, Carlos H.; Stoica, Adrian; Cwik, Thomas

    2000-01-01

    The quantum mechanical functionality of nanoelectronic devices such as resonant tunneling diodes (RTDs), quantum well infrared-photodetectors (QWIPs), quantum well lasers, and heterostructure field effect transistors (HFETs) is enabled by material variations on an atomic scale. The design and optimization of such devices requires a fundamental understanding of electron transport in such dimensions. The Nanoelectronic Modeling Tool (NEMO) is a general-purpose quantum device design and analysis tool based on a fundamental non-equilibrium electron transport theory. NEW was combined with a parallelized genetic algorithm package (PGAPACK) to evolve structural and material parameters to match a desired set of experimental data. A numerical experiment that evolves structural variations such as layer widths and doping concentrations is performed to analyze an experimental current voltage characteristic. The genetic algorithm is found to drive the NEMO simulation parameters close to the experimentally prescribed layer thicknesses and doping profiles. With such a quantitative agreement between theory and experiment design synthesis can be performed.

  16. Genetic Engineering and Crop Production.

    Science.gov (United States)

    Jones, Helen C.; Frost, S.

    1991-01-01

    With a spotlight upon current agricultural difficulties and environmental dilemmas, this paper considers both the extant and potential applications of genetic engineering with respect to crop production. The nonagricultural factors most likely to sway the impact of this emergent technology upon future crop production are illustrated. (JJK)

  17. Genetic Engineering and Crop Production.

    Science.gov (United States)

    Jones, Helen C.; Frost, S.

    1991-01-01

    With a spotlight upon current agricultural difficulties and environmental dilemmas, this paper considers both the extant and potential applications of genetic engineering with respect to crop production. The nonagricultural factors most likely to sway the impact of this emergent technology upon future crop production are illustrated. (JJK)

  18. Mesenchymal stem cells support hepatocyte function in engineered liver grafts.

    Science.gov (United States)

    Kadota, Yoshie; Yagi, Hiroshi; Inomata, Kenta; Matsubara, Kentaro; Hibi, Taizo; Abe, Yuta; Kitago, Minoru; Shinoda, Masahiro; Obara, Hideaki; Itano, Osamu; Kitagawa, Yuko

    2014-01-01

    Recent studies suggest that organ decellularization is a promising approach to facilitate the clinical application of regenerative therapy by providing a platform for organ engineering. This unique strategy uses native matrices to act as a reservoir for the functional cells which may show therapeutic potential when implanted into the body. Appropriate cell sources for artificial livers have been debated for some time. The desired cell type in artificial livers is primary hepatocytes, but in addition, other supportive cells may facilitate this stem cell technology. In this context, the use of mesenchymal stem cells (MSC) is an option meeting the criteria for therapeutic organ engineering. Ideally, supportive cells are required to (1) reduce the hepatic cell mass needed in an engineered liver by enhancing hepatocyte function, (2) modulate hepatic regeneration in a paracrine fashion or by direct contact, and (3) enhance the preservability of parenchymal cells during storage. Here, we describe enhanced hepatic function achieved using a strategy of sequential infusion of cells and illustrate the advantages of co-cultivating bone marrow-derived MSCs with primary hepatocytes in the engineered whole-liver scaffold. These co-recellularized liver scaffolds colonized by MSCs and hepatocytes were transplanted into live animals. After blood flow was established, we show that expression of adhesion molecules and proangiogenic factors was upregulated in the graft.

  19. Musculoskeletal tissue engineering with human umbilical cord mesenchymal stromal cells

    Science.gov (United States)

    Wang, Limin; Ott, Lindsey; Seshareddy, Kiran; Weiss, Mark L; Detamore, Michael S

    2011-01-01

    Multipotent mesenchymal stromal cells (MSCs) hold tremendous promise for tissue engineering and regenerative medicine, yet with so many sources of MSCs, what are the primary criteria for selecting leading candidates? Ideally, the cells will be multipotent, inexpensive, lack donor site morbidity, donor materials should be readily available in large numbers, immunocompatible, politically benign and expandable in vitro for several passages. Bone marrow MSCs do not meet all of these criteria and neither do embryonic stem cells. However, a promising new cell source is emerging in tissue engineering that appears to meet these criteria: MSCs derived from Wharton’s jelly of umbilical cord MSCs. Exposed to appropriate conditions, umbilical cord MSCs can differentiate in vitro along several cell lineages such as the chondrocyte, osteoblast, adipocyte, myocyte, neuronal, pancreatic or hepatocyte lineages. In animal models, umbilical cord MSCs have demonstrated in vivo differentiation ability and promising immunocompatibility with host organs/tissues, even in xenotransplantation. In this article, we address their cellular characteristics, multipotent differentiation ability and potential for tissue engineering with an emphasis on musculoskeletal tissue engineering. PMID:21175290

  20. Non-viral approaches for direct conversion into mesenchymal cell types: Potential application in tissue engineering.

    Science.gov (United States)

    Lee, Eun-Seo; Kim, Seung Hyun L; Lee, Hwajin; Hwang, Nathaniel S

    2016-05-01

    Acquiring adequate number of cells is one of the crucial factors to apply tissue engineering strategies in order to recover critical-sized defects. While the reprogramming technology used for inducing pluripotent stem cells (iPSCs) opened up a direct path for generating pluripotent stem cells, a direct conversion strategy may provide another possibility to obtain desired cells for tissue engineering. In order to convert a somatic cell into any other cell type, diverse approaches have been investigated. Conspicuously, in contrast to traditional viral transduction method, non-viral delivery of conversion factors has the merit of lowering immune responses and provides safer genetic manipulation, thus revolutionizing the generation of directly converted cells and its application in therapeutics. In addition, applying various microenvironmental modulations have potential to ameliorate the conversion of somatic cells into different lineages. In this review, we discuss the recent progress in direct conversion technologies, specifically focusing on generating mesenchymal cell types. © 2016 Wiley Periodicals, Inc.

  1. Development of 3D in vitro platform technology to engineer mesenchymal stem cells

    Directory of Open Access Journals (Sweden)

    Hosseinkhani H

    2012-06-01

    Full Text Available Hossein Hosseinkhani,1 Po-Da Hong,1 Dah-Shyong Yu,2 Yi-Ru Chen,3 Diana Ickowicz,4 Ira-Yudovin Farber,4 Abraham J Domb41Graduate Institute of Biomedical Engineering, National Taiwan University of Science and Technology (TAIWANTECH, 2Nanomedicine Research Center, National Defense Medical Center, Taipei, Taiwan, 3Department of Biomedical Engineering, National Yang-Ming University, Taipei, Taiwan, 4Institute of Drug Research, The Center for Nanoscience and Nanotechnology, School of Pharmacy-Faculty of Medicine, The Hebrew University of Jerusalem, Jerusalem, IsraelAbstract: This study aims to develop a three-dimensional in vitro culture system to genetically engineer mesenchymal stem cells (MSC to express bone morphogenic protein-2. We employed nanofabrication technologies borrowed from the spinning industry, such as electrospinning, to mass-produce identical building blocks in a variety of shapes and sizes to fabricate electrospun nanofiber sheets comprised of composites of poly (glycolic acid and collagen. Homogenous nanoparticles of cationic biodegradable natural polymer were formed by simple mixing of an aqueous solution of plasmid DNA encoded bone morphogenic protein-2 with the same volume of cationic polysaccharide, dextran-spermine. Rat bone marrow MSC were cultured on electrospun nanofiber sheets comprised of composites of poly (glycolic acid and collagen prior to the incorporation of the nanoparticles into the nanofiber sheets. Bone morphogenic protein-2 was significantly detected in MSC cultured on nanofiber sheets incorporated with nanoparticles after 2 days compared with MSC cultured on nanofiber sheets incorporated with naked plasmid DNA. We conclude that the incorporation of nanoparticles into nanofiber sheets is a very promising strategy to genetically engineer MSC and can be used for further applications in regenerative medicine therapy.Keywords: 3D culture, nanoparticles, nanofibers, polycations, tissue engineering

  2. Overcoming Challenges in Engineering the Genetic Code.

    Science.gov (United States)

    Lajoie, M J; Söll, D; Church, G M

    2016-02-27

    Withstanding 3.5 billion years of genetic drift, the canonical genetic code remains such a fundamental foundation for the complexity of life that it is highly conserved across all three phylogenetic domains. Genome engineering technologies are now making it possible to rationally change the genetic code, offering resistance to viruses, genetic isolation from horizontal gene transfer, and prevention of environmental escape by genetically modified organisms. We discuss the biochemical, genetic, and technological challenges that must be overcome in order to engineer the genetic code.

  3. Tooth engineering: searching for dental mesenchymal cells sources.

    Directory of Open Access Journals (Sweden)

    Laetitia eKeller

    2011-03-01

    Full Text Available The implantation of cultured re-associations between embryonic dental mesenchymal cells and epithelial cells from mouse molars at ED14 allowed making full teeth with crown, root, periodontal ligament fibers and bone. Although representing valuable tools to set up methodologies embryonic cells are not easily available. This work thus aimed to replace the embryonic cells by dental mesenchymal cell lines or cultured expanded embryonic cells, and to test their ability to mediate tooth development in vitro when re-associated with a competent dental epithelium. Histology, immunostaining and RT-PCR allowed getting complementary sets of results. Two different immortalized cell lines from ED18 dental mesenchyme failed in mediating tooth formation. The potentialities of embryonic dental mesenchymal cells decreased from ED14 to ED16 and were lost at ED18. This is likely related to a change in the mesenchymal cell phenotype and/or populations during development. Attempts to cultivate ED14 or ED16 embryonic dental mesenchymal cells prior to re-association led to the loss of their ability to support tooth development. This was accompanied by a down-regulation of Fgf3 transcription. Supplementation of the culture medium with FGF2 allowed restoring Fgf3 expression, but not the ability of mesenchymal cells to engage in tooth formation. Altogether, these observations suggest that a competent cell population exists in the dental mesenchyme at ED14, progressively decreases during development, and cannot as such be maintained in vitro. This study evidenced the need for specific conditions to maintain the ability of dental mesenchymal cells to initiate whole tooth formation, when re-associated with an odontogenic epithelium. Efforts to improve the culture conditions will have to be combined with attempts to characterize the competent cells within the dental mesenchyme.

  4. Genetically Engineered Immunotherapy for Advanced Cancer

    Science.gov (United States)

    In this trial, doctors will collect T lymphocytes from patients with advanced mesothelin-expressing cancer and genetically engineer them to recognize mesothelin. The gene-engineered cells will be multiplied and infused into the patient to fight the cancer

  5. Development of 3D in vitro platform technology to engineer mesenchymal stem cells.

    Science.gov (United States)

    Hosseinkhani, Hossein; Hong, Po-Da; Yu, Dah-Shyong; Chen, Yi-Ru; Ickowicz, Diana; Farber, Ira-Yudovin; Domb, Abraham J

    2012-01-01

    This study aims to develop a three-dimensional in vitro culture system to genetically engineer mesenchymal stem cells (MSC) to express bone morphogenic protein-2. We employed nanofabrication technologies borrowed from the spinning industry, such as electrospinning, to mass-produce identical building blocks in a variety of shapes and sizes to fabricate electrospun nanofiber sheets comprised of composites of poly (glycolic acid) and collagen. Homogenous nanoparticles of cationic biodegradable natural polymer were formed by simple mixing of an aqueous solution of plasmid DNA encoded bone morphogenic protein-2 with the same volume of cationic polysaccharide, dextran-spermine. Rat bone marrow MSC were cultured on electrospun nanofiber sheets comprised of composites of poly (glycolic acid) and collagen prior to the incorporation of the nanoparticles into the nanofiber sheets. Bone morphogenic protein-2 was significantly detected in MSC cultured on nanofiber sheets incorporated with nanoparticles after 2 days compared with MSC cultured on nanofiber sheets incorporated with naked plasmid DNA. We conclude that the incorporation of nanoparticles into nanofiber sheets is a very promising strategy to genetically engineer MSC and can be used for further applications in regenerative medicine therapy.

  6. Genetic engineering and coagulation factors.

    Science.gov (United States)

    Fass, D N; Toole, J J

    1985-06-01

    It is unfortunate that we cannot report, in the area of coagulation, advances that have been seen in related fields such as thrombolytic therapy. The reported progress (Gold et al, 1984; Van de Werf et al, 1984) with human recombinant tissue plasminogen activator (Pennica et al, 1983) augers well for the application of recombinant technology to the problems faced by patients with coagulation defects. While plasminogen activator is being assessed in an acute therapeutic setting, its use signals a beginning of the application of the technology to abnormalities of the haemostatic mechanism. Chronic administration of coagulation factors for prophylaxis and replacement therapy would appear to be just one more step down the pathway illuminated by the biochemists, microbiologists and cell biologists who have preceded the clinicians in this promising area. There is no record of the use of genetically engineered materials in the treatment of coagulation defects, primarily because the body of knowledge and refined techniques have only recently been acquired. For this reason we have had to project developments in other areas onto the problems that exist for the haemostatically compromised patient. In describing the potential usefulness of these technologies, it is difficult to ascertain where the logical projection, from a fully investigated model system, diverges from flights of imaginative fancy. Cloning projects considered overly ambitious and grandiose at the beginning of this decade are already accomplished feats. The feasibility of gene therapy in the mammalian system has been demonstrated, and trade publications now discuss governmental approval for investigative use of this procedure in 1985. Panels of physicians, scientists and even politicians now seriously contemplate and promulgate views and regulations pertaining to the efficacy and ethics of the use of genetic engineering in the treatment of human disease. The haemophilias will certainly be among the first

  7. Genetic Engineering of Alfalfa (Medicago sativa L.).

    Science.gov (United States)

    Wang, Dan; Khurshid, Muhammad; Sun, Zhan Min; Tang, Yi Xiong; Zhou, Mei Liang; Wu, Yan Min

    2016-01-01

    Alfalfa is excellent perennial legume forage for its extensive ecological adaptability, high nutrition value, palatability and biological nitrogen fixation. It plays a very important role in the agriculture, animal husbandry and ecological construction. It is cultivated in all continents. With the development of modern plant breeding and genetic engineering techniques, a large amount of work has been carried out on alfalfa. Here we summarize the recent research advances in genetic engineering of alfalfa breeding, including transformation, quality improvement, stress resistance and as a bioreactor. The review article can enables us to understand the research method, direction and achievements of genetic engineering technology of Alfalfa.

  8. Targeted Eradication of Prostate Cancer Mediated by Engineered Mesenchymal Stem

    Science.gov (United States)

    2007-12-01

    mesenchymal tissues , such as bone, cartilage, adipose and connective tissues . Early studies demonstrated that mouse marrow stroma cell line...CBFV) using transcranial Doppler sonography (TCD), together with additional indices including salivary cortisol and subjective state. Two studies at...detrimental side effects to the patients. Even though the exact mechanism of preferential prostate cancer bone metastasis has not yet been well

  9. Optimizing viral and non-viral gene transfer methods for genetic modification of porcine mesenchymal stem cells

    DEFF Research Database (Denmark)

    Stiehler, Maik; Duch, Mogens R.; Mygind, Tina

    2006-01-01

    INTRODUCTION: Mesenchymal stem cells (MSCs) provide an excellent source of pluripotent progenitor cells for tissue-engineering applications due to their proliferation capacity and differentiation potential. Genetic modification of MSCs with genes encoding tissue-specific growth factors and cytoki......INTRODUCTION: Mesenchymal stem cells (MSCs) provide an excellent source of pluripotent progenitor cells for tissue-engineering applications due to their proliferation capacity and differentiation potential. Genetic modification of MSCs with genes encoding tissue-specific growth factors...... viral and non-viral ex vivo gene delivery systems with respect to gene transfer efficiency, maintenance of transgene expression, and safety issues using primary porcine MSCs as target cells. MATERIALS AND METHODS: MSCs were purified from bone marrow aspirates from the proximal tibiae of four 3-month......-old Danish landrace pigs by Ficoll step gradient separation and polystyrene adherence technique. Vectors expressing enhanced green fluorescent protein (eGFP) and human bone morphogenetic protein-2 (BMP-2) were transferred to the cells by different non-viral methods and by use of recombinant adeno...

  10. PROSPECTS FOR APPLICATION OF Aplysinidae FAMILY MARINE SPONGE SKELETONS AND MESENCHYMAL STROMAL CELLS IN TISSUE ENGINEERING

    Directory of Open Access Journals (Sweden)

    О. Yu. Rogulska

    2011-10-01

    Full Text Available Development of the new types of tissue engineered structures is one of the promising trends of current biotechnology. The study was directed to the assessment of prospects for the application of chitin-based skeletons derived from marine sponges of Aplysinidae family (Aplysina fulva and Aplysina aerophoba for creation of bioengineered constructs based on human mesenchymal stromal cells. After cleaning and demineralization procedures, sponge skeletons appeared as three-dimensional macroporous matrices formed by intersecting chitin fibrils. After seeding into chitin-based matrices the cells were attached to the surface of the fibrils and were able to spread and proliferate. Mesenchymal stromal cells within Aplysina fulva differentiated into osteogenic and adipogenic directions under the influence of appropriate inductors. Demineralized skeletons derived from marine sponges of Aplysinidae family could be used as scaffolds for mesenchymal stromal cells which provides new opportunities for the creation of adipose and bone tissue engineered structures.

  11. Recent Advances in Genetic Engineering - A Review

    OpenAIRE

    Sobiah Rauf; Zubair Anwar; Hussain Mustatab Wahedi; Jabar Zaman Khan Khattak; Talal Jamil

    2012-01-01

    Humans have been doing genetic engineering, a technology which is transforming our world, for thousands of years on a wide range of plants, animals and micro organism and have applications in the field of medicine, research, industry and agriculture. The rapid developments in the field of genetic engineering have given a new impetus to biotechnology. This introduces the possibility of tailoring organisms in order to optimize the production of established or novel metabolites of commercial imp...

  12. Genetic elements of plant viruses as tools for genetic engineering.

    OpenAIRE

    Mushegian, A R; Shepherd, R J

    1995-01-01

    Viruses have developed successful strategies for propagation at the expense of their host cells. Efficient gene expression, genome multiplication, and invasion of the host are enabled by virus-encoded genetic elements, many of which are well characterized. Sequences derived from plant DNA and RNA viruses can be used to control expression of other genes in vivo. The main groups of plant virus genetic elements useful in genetic engineering are reviewed, including the signals for DNA-dependent a...

  13. Natural genetic engineering: intelligence & design in evolution?

    DEFF Research Database (Denmark)

    Ussery, David

    2011-01-01

    function. Shapiro argues that what we see in genomes is 'Natural Genetic Engineering', or designed evolution: "Thinking about genomes from an informatics perspective, it is apparent that systems engineering is a better metaphor for the evolutionary process than the conventional view of evolution...

  14. 130 FEMINISM AND HUMAN GENETIC ENGINEERING: A ...

    African Journals Online (AJOL)

    Ike Odimegwu

    Abstract. Human genetic in the area of Bio-ethics is a new, rapidly advancing. Science. ... Human genetic engineering, a recent one in medical science and practice, is one ..... The Church on Cloning and Stem Cell Research. The teaching of ...

  15. Genetic Engineering: The Modification of Man

    Science.gov (United States)

    Sinsheimer, Robert L.

    1970-01-01

    Describes somatic and genetic manipulations of individual genotypes, using diabetes control as an example of the first mode that is potentially realizable be derepression or viral transduction of genes. Advocates the use of genetic engineering of the second mode to remove man from his biological limitations, but offers maxims to ensure the…

  16. Controlled Inhibition of the Mesenchymal Stromal Cell Pro-inflammatory Secretome via Microparticle Engineering

    Directory of Open Access Journals (Sweden)

    Sudhir H. Ranganath

    2016-06-01

    Full Text Available Mesenchymal stromal cells (MSCs are promising therapeutic candidates given their potent immunomodulatory and anti-inflammatory secretome. However, controlling the MSC secretome post-transplantation is considered a major challenge that hinders their clinical efficacy. To address this, we used a microparticle-based engineering approach to non-genetically modulate pro-inflammatory pathways in human MSCs (hMSCs under simulated inflammatory conditions. Here we show that microparticles loaded with TPCA-1, a small-molecule NF-κB inhibitor, when delivered to hMSCs can attenuate secretion of pro-inflammatory factors for at least 6 days in vitro. Conditioned medium (CM derived from TPCA-1-loaded hMSCs also showed reduced ability to attract human monocytes and prevented differentiation of human cardiac fibroblasts to myofibroblasts, compared with CM from untreated or TPCA-1-preconditioned hMSCs. Thus, we provide a broadly applicable bioengineering solution to facilitate intracellular sustained release of agents that modulate signaling. We propose that this approach could be harnessed to improve control over MSC secretome post-transplantation, especially to prevent adverse remodeling post-myocardial infarction.

  17. Genetic engineering of microbial pesticides

    Science.gov (United States)

    Bruce C. Carlton

    1985-01-01

    Recent advances in genetics and molecular biology make possible the cloning and genetic manipulation of genes for insecticidal activities from natural insect pathogens. Using recombinant DNA methods and site-directed mutagenesis of specific gene regions, production of new and improved biorationals should be possible.

  18. Commercialising genetically engineered animal biomedical products.

    Science.gov (United States)

    Sullivan, Eddie J; Pommer, Jerry; Robl, James M

    2008-01-01

    Research over the past two decades has increased the quality and quantity of tools available to produce genetically engineered animals. The number of potentially viable biomedical products from genetically engineered animals is increasing. However, moving from cutting-edge research to development and commercialisation of a biomedical product that is useful and wanted by the public has significant challenges. Even early stage development of genetically engineered animal applications requires consideration of many steps, including quality assurance and quality control, risk management, gap analysis, founder animal establishment, cell banking, sourcing of animals and animal-derived material, animal facilities, product collection facilities and processing facilities. These steps are complicated and expensive. Biomedical applications of genetically engineered animals have had some recent successes and many applications are well into development. As researchers consider applications for their findings, having a realistic understanding of the steps involved in the development and commercialisation of a product, produced in genetically engineered animals, is useful in determining the risk of genetic modification to the animal nu. the potential public benefit of the application.

  19. Recent Advances in Genetic Engineering - A Review

    Directory of Open Access Journals (Sweden)

    Sobiah Rauf

    2012-01-01

    Full Text Available Humans have been doing genetic engineering, a technology which is transforming our world, for thousands of years on a wide range of plants, animals and micro organism and have applications in the field of medicine, research, industry and agriculture. The rapid developments in the field of genetic engineering have given a new impetus to biotechnology. This introduces the possibility of tailoring organisms in order to optimize the production of established or novel metabolites of commercial importance and of transferring genetic material from one organism to another. In order to achieve potential benefits of genetic engineering the only need is to develop perfect tools and techniques. Once it has been perfected then all of the problems associated with food production can be solved, the world environment can be restored, and human health and lifestyle will improve beyond imagination. No doubt that there are almost no limits to what can be achieved through responsible genetic engineering. Classical field of genetic engineering and some of its advancements are discussed in this review.

  20. Encapsulation of factor IX-engineered mesenchymal stem cells in fibrinogen-alginate microcapsules enhances their viability and transgene secretion.

    Science.gov (United States)

    Sayyar, Bahareh; Dodd, Megan; Wen, Jianping; Ma, Shirley; Marquez-Curtis, Leah; Janowska-Wieczorek, Anna; Hortelano, Gonzalo

    2012-01-01

    Cell microencapsulation holds significant promise as a strategy for cellular therapies; however, inadequate survival and functionality of the enclosed cells limit its application in hemophilia treatment. Here, we evaluated the use of alginate-based microcapsules to enhance the viability and transgene secretion of human cord blood-derived mesenchymal stem cells in three-dimensional cultures. Given the positive effects of extracellular matrix molecules on mesenchymal stem cell growth, we tested whether fibrinogen-supplemented alginate microcapsules can improve the efficiency of encapsulated factor IX-engineered mesenchymal stem cells as a treatment of hemophilia B. We found that fibrinogen-supplemented alginate microcapsules (a) significantly enhanced the viability and proliferation of factor IX-engineered mesenchymal stem cells and (b) increased factor IX secretion by mesenchymal stem cells compared to mesenchymal stem cells in nonsupplemented microcapsules. Moreover, we observed the osteogenic, but not chondrogenic or adipogenic, differentiation capability of factor IX-engineered cord blood mesenchymal stem cells and their efficient factor IX secretion while encapsulated in fibrinogen-supplemented alginate microcapsules. Thus, the use of engineered mesenchymal stem cells encapsulated in fibrinogen-modified microcapsules may have potential application in the treatment of hemophilia or other protein deficiency diseases.

  1. Encapsulation of factor IX–engineered mesenchymal stem cells in fibrinogen–alginate microcapsules enhances their viability and transgene secretion

    Directory of Open Access Journals (Sweden)

    Bahareh Sayyar

    2012-12-01

    Full Text Available Cell microencapsulation holds significant promise as a strategy for cellular therapies; however, inadequate survival and functionality of the enclosed cells limit its application in hemophilia treatment. Here, we evaluated the use of alginate-based microcapsules to enhance the viability and transgene secretion of human cord blood–derived mesenchymal stem cells in three-dimensional cultures. Given the positive effects of extracellular matrix molecules on mesenchymal stem cell growth, we tested whether fibrinogen-supplemented alginate microcapsules can improve the efficiency of encapsulated factor IX–engineered mesenchymal stem cells as a treatment of hemophilia B. We found that fibrinogen-supplemented alginate microcapsules (a significantly enhanced the viability and proliferation of factor IX–engineered mesenchymal stem cells and (b increased factor IX secretion by mesenchymal stem cells compared to mesenchymal stem cells in nonsupplemented microcapsules. Moreover, we observed the osteogenic, but not chondrogenic or adipogenic, differentiation capability of factor IX–engineered cord blood mesenchymal stem cells and their efficient factor IX secretion while encapsulated in fibrinogen-supplemented alginate microcapsules. Thus, the use of engineered mesenchymal stem cells encapsulated in fibrinogen-modified microcapsules may have potential application in the treatment of hemophilia or other protein deficiency diseases.

  2. Advances in genetic engineering of domestic animals

    Directory of Open Access Journals (Sweden)

    Shaohua WANG,Kun ZHANG,Yunping DAI

    2016-03-01

    Full Text Available Global population will increase to over nine billion by 2050 with the doubling in demand for meat and milk. To overcome this challenge, it is necessary to breed highly efficient and productive livestock. Furthermore, livestock are also excellent models for human diseases and ideal bioreactors to produce pharmaceutical proteins. Thus, genetic engineering of domestic animals presents a critical and valuable tool to address these agricultural and biomedical applications. Overall, genetic engineering has evolved through three stages in history: transgenesis, gene targeting, and gene editing. Since the birth of the first transgenic pig, genetic engineering in livestock has been advancing slowly due to inherent technical limitations. A major breakthrough has been the advent of somatic cell nuclear transfer, which, for the first time, provided the technical ability to produce site-specific genome-modified domestic animals. However, the low efficiency of gene targeting events in somatic cells prohibits its wide use in agricultural and biomedical applications. Recently, rapid progress in tools and methods of genome engineering has been made, allowing genetic editing from mutation of a single base pair to the deletion of entire chromosomes. Here, we review the major advances of genetic engineering in domestic animals with emphasis placed on the introduction of latest designer nucleases.

  3. Genetically engineered nanocarriers for drug delivery

    Directory of Open Access Journals (Sweden)

    Shi P

    2014-03-01

    Full Text Available Pu Shi, Joshua A Gustafson, J Andrew MacKayDepartment of Pharmacology and Pharmaceutical Sciences, University of Southern California, Los Angeles, CA, USAAbstract: Cytotoxicity, low water solubility, rapid clearance from circulation, and off-target side-effects are common drawbacks of conventional small-molecule drugs. To overcome these shortcomings, many multifunctional nanocarriers have been proposed to enhance drug delivery. In concept, multifunctional nanoparticles might carry multiple agents, control release rate, biodegrade, and utilize target-mediated drug delivery; however, the design of these particles presents many challenges at the stage of pharmaceutical development. An emerging solution to improve control over these particles is to turn to genetic engineering. Genetically engineered nanocarriers are precisely controlled in size and structure and can provide specific control over sites for chemical attachment of drugs. Genetically engineered drug carriers that assemble nanostructures including nanoparticles and nanofibers can be polymeric or non-polymeric. This review summarizes the recent development of applications in drug and gene delivery utilizing nanostructures of polymeric genetically engineered drug carriers such as elastin-like polypeptides, silk-like polypeptides, and silk-elastin-like protein polymers, and non-polymeric genetically engineered drug carriers such as vault proteins and viral proteins.Keywords: polymeric drug carrier, non-polymeric drug carrier, gene delivery, GE drug carriers

  4. Genetic Engineering Strategies for Enhanced Biodiesel Production.

    Science.gov (United States)

    Hegde, Krishnamoorthy; Chandra, Niharika; Sarma, Saurabh Jyoti; Brar, Satinder Kaur; Veeranki, Venkata Dasu

    2015-07-01

    The focus on biodiesel research has shown a tremendous growth over the last few years. Several microbial and plant sources are being explored for the sustainable biodiesel production to replace the petroleum diesel. Conventional methods of biodiesel production have several limitations related to yield and quality, which led to development of new engineering strategies to improve the biodiesel production in plants, and microorganisms. Substantial progress in utilizing algae, yeast, and Escherichia coli for the renewable production of biodiesel feedstock via genetic engineering of fatty acid metabolic pathways has been reported in the past few years. However, in most of the cases, the successful commercialization of such engineering strategies for sustainable biodiesel production is yet to be seen. This paper systematically presents the drawbacks in the conventional methods for biodiesel production and an exhaustive review on the present status of research in genetic engineering strategies for production of biodiesel in plants, and microorganisms. Further, we summarize the technical challenges need to be tackled to make genetic engineering technology economically sustainable. Finally, the need and prospects of genetic engineering technology for the sustainable biodiesel production and the recommendations for the future research are discussed.

  5. Engineered three-dimensional rabbit oral epithelial-mesenchymal-muscular hybrid sheets

    Institute of Scientific and Technical Information of China (English)

    Shigeki Yamane; Kazunari Higa; Takashi Umezawa; Masamitsu Serikawa; Jun Shimazaki; Shinichi Abe

    2016-01-01

    Regenerative muscles are required for swallowing and mastication, and are important for functional recovery from diseases involving oral muscular defects. Therefore, we generated three-layer hybrid sheets, similar to oral mucosal structures containing submucosal muscles, using rabbit oral mucosa epithelial, mesenchymal, and myoblastic progenitor cells, and examined the structural proteins. Each cell type was obtained from rabbit oral mucosa using enzymatic digestion. Isolated mesenchymal and myoblastic cells were multi-differentiated into osteoblasts, adipocytes, and chondrocytes or myotubes. Isolated epithelial cells were cultured on collagen gels containing isolated mesenchymal cells for 2 weeks, and these epithelial–mesenchymal cell sheets were laminated onto myoblastic cell sheets. The engineered hybrid sheets were multi-stratified in the epithelial and myoblastic layers in a time-dependent manner, expressing intermediate cytoskeletal filament proteins of epithelium and muscle. Hybrid sheets also expressed extracellular matrix basement membrane proteins. Immature cell markers for epithelial and myoblastic cells were observed continuously in hybrid sheet cultures. We established engineered three-dimensional rabbit oral mucosa hybrid sheets containing each immature cell type in vitro.

  6. Genetic Engineering: and the Law

    Science.gov (United States)

    Australian Journal of Mental Retardation, 1977

    1977-01-01

    In a transcript from a radio show, Nobel Prize Winner Sir Macfarlane Burnet stresses the critical need for scientists to regulate their own activities in genetic research and cites the potential danger of creating a new form of polio which might escape. (CL)

  7. Genetic engineering for skeletal regenerative medicine.

    Science.gov (United States)

    Gersbach, Charles A; Phillips, Jennifer E; García, Andrés J

    2007-01-01

    The clinical challenges of skeletal regenerative medicine have motivated significant advances in cellular and tissue engineering in recent years. In particular, advances in molecular biology have provided the tools necessary for the design of gene-based strategies for skeletal tissue repair. Consequently, genetic engineering has emerged as a promising method to address the need for sustained and robust cellular differentiation and extracellular matrix production. As a result, gene therapy has been established as a conventional approach to enhance cellular activities for skeletal tissue repair. Recent literature clearly demonstrates that genetic engineering is a principal factor in constructing effective methods for tissue engineering approaches to bone, cartilage, and connective tissue regeneration. This review highlights this literature, including advances in the development of efficacious gene carriers, novel cell sources, successful delivery strategies, and optimal target genes. The current status of the field and the challenges impeding the clinical realization of these approaches are also discussed.

  8. Advances in genetic engineering of domestic animals

    OpenAIRE

    Shaohua WANG,Kun ZHANG,Yunping DAI

    2016-01-01

    Global population will increase to over nine billion by 2050 with the doubling in demand for meat and milk. To overcome this challenge, it is necessary to breed highly efficient and productive livestock. Furthermore, livestock are also excellent models for human diseases and ideal bioreactors to produce pharmaceutical proteins. Thus, genetic engineering of domestic animals presents a critical and valuable tool to address these agricultural and biomedical applications. Overall, genetic enginee...

  9. Advances in genetic engineering of domestic animals

    OpenAIRE

    Shaohua WANG,Kun ZHANG,Yunping DAI

    2016-01-01

    Global population will increase to over nine billion by 2050 with the doubling in demand for meat and milk. To overcome this challenge, it is necessary to breed highly efficient and productive livestock. Furthermore, livestock are also excellent models for human diseases and ideal bioreactors to produce pharmaceutical proteins. Thus, genetic engineering of domestic animals presents a critical and valuable tool to address these agricultural and biomedical applications. Overall, genetic enginee...

  10. Genetically Engineered Crops: Experiences and Prospects

    NARCIS (Netherlands)

    Giller, K.E.

    2016-01-01

    Since their introduction in the mid-1990s, genetically engineered (GE) crops have been the topic of much debate. This report reviews evidence accumulated from experiences on the most widely grown GE crops to date: herbicide-resistant and insect-resistant varieties of maize, soybean, and cotton. Whil

  11. Genetically Engineered Crops: Experiences and Prospects

    NARCIS (Netherlands)

    Giller, K.E.

    2016-01-01

    Since their introduction in the mid-1990s, genetically engineered (GE) crops have been the topic of much debate. This report reviews evidence accumulated from experiences on the most widely grown GE crops to date: herbicide-resistant and insect-resistant varieties of maize, soybean, and cotton.

  12. [The microencapsulated genetic engineering cells: a new platform on treatment of cancer instead of genetic engineering drugs].

    Science.gov (United States)

    Pan, Yuelong; Zheng, Shu

    2003-06-01

    The microencapsulated genetic cells may be a new platform instead of genetic engineering drugs, as they can overcome the genetic engineering drugs' shortages such as short half-life in vivo, low activity, and incomplete elimination of organic solvent. This article reviews and summarizes the advantages, possible problems and solution and the feasibility of using microencapsulated genetic engineering cells in the treatment of cancer.

  13. Maintenance of bladder urothelia integrity and successful urothelialization of various tissue-engineered mesenchymes in vitro.

    Science.gov (United States)

    Bouhout, Sara; Tremblay, Julie; Bolduc, Stephane

    2015-10-01

    Tissue-engineering offers the opportunity to produce hybrid tissues in vitro. The induction of bladder urothelial cells (BUCs) differentiation in vitro has been assessed by several research groups to build bladder models for fundamental studies and clinical applications. However, BUC induction of advanced differentiation in culture remains a challenging task. To reach this goal, optimal culture conditions are required, notably the use of specific additives as well as proper mesenchymal support. The best positive control for BUCs functional state monitoring is native urothelium collected from healthy bladder samples. In order to establish the best culture conditions to maintain and promote BUC differentiated state, native urothelia were cultured on various mesenchymes. Native bladder mesenchymes were used as controls for the maintenance of native urothelia. Histological and ultrastructural analyses showed the necessity to have a cellularized mesenchyme for rapid formation of a pseudostratified urothelium, allowing apical membrane rearrangement of the superficial cells in culture. Taken together, the results strongly suggest that it is possible to conserve the integrity of urothelia in vitro and, thus, potentially use them for eventual clinical applications and pharmacological investigations.

  14. Chromosome engineering: power tools for plant genetics.

    Science.gov (United States)

    Chan, Simon W L

    2010-12-01

    The term "chromosome engineering" describes technologies in which chromosomes are manipulated to change their mode of genetic inheritance. This review examines recent innovations in chromosome engineering that promise to greatly increase the efficiency of plant breeding. Haploid Arabidopsis thaliana have been produced by altering the kinetochore protein CENH3, yielding instant homozygous lines. Haploid production will facilitate reverse breeding, a method that downregulates recombination to ensure progeny contain intact parental chromosomes. Another chromosome engineering success is the conversion of meiosis into mitosis, which produces diploid gametes that are clones of the parent plant. This is a key step in apomixis (asexual reproduction through seeds) and could help to preserve hybrid vigor in the future. New homologous recombination methods in plants will potentiate many chromosome engineering applications. Copyright © 2010 Elsevier Ltd. All rights reserved.

  15. What Ideas Do Students Associate with "Biotechnology" and "Genetic Engineering"?

    Science.gov (United States)

    Hill, Ruaraidh; Stanisstreet, Martin; Boyes, Edward

    2000-01-01

    Explores the ideas that students aged 16-19 associate with the terms 'biotechnology' and 'genetic engineering'. Indicates that some students see biotechnology as risky whereas genetic engineering was described as ethically wrong. (Author/ASK)

  16. Hypoxia and Stem Cell-Based Engineering of Mesenchymal Tissues

    OpenAIRE

    Ma, Teng; Grayson, Warren L.; Fröhlich, Mirjam; Vunjak-Novakovic, Gordana

    2009-01-01

    Stem cells have the ability for prolonged self-renewal and differentiation into mature cells of various lineages, which makes them important cell sources for tissue engineering applications. Their remarkable ability to replenish and differentiate in vivo is regulated by both intrinsic and extrinsic cellular mechanisms. The anatomical location where the stem cells reside, known as the “stem cell niche or microenvironment,” provides signals conducive to the maintenance of definitive stem cell p...

  17. Mesenchymal stem cells from the oral cavity and their potential value in tissue engineering.

    Science.gov (United States)

    Sanz, Antonio R; Carrión, Flavio S; Chaparro, Alejandra P

    2015-02-01

    Periodontal disease is one of the most common conditions affecting humans, and current treatment strategies, which focus on the removal and long-term control of dental plaque, are generally successful in eliminating active disease and promoting tissue repair. However, regeneration of the supporting structures of the tooth remains an elusive goal and a challenge. The formation of new bone and cementum with supportive periodontal ligament is the ultimate objective, but current regeneration therapies are incapable of achieving this in a predictable way. The regeneration of periodontal tissue requires a combination of fundamental events, such as appropriate level and sequencing of regulatory signals, the presence of progenitor cells, an extracellular matrix or carrier and an adequate blood supply. Based on tissue-engineering concepts, the regeneration process may be modulated by manipulating the signaling pathways of regulatory molecules, the extracellular matrix or scaffold, or the cellular components. The identification of mesenchymal stem cells from bone marrow started a new era in regenerative medicine. Tissue engineering using mesenchymal stem cells became a therapeutic option with several advantages, including high-quality regeneration of damaged tissues without the formation of fibrous tissue, minimal donor-site morbidity compared with autografts and a low risk of autoimmune rejection and disease transmission. The aim of this review was to describe the main sources of mesenchymal stem cells from tissues in the oral cavity and the potential of these cells in regenerative therapy. Special attention is paid to gingival tissue-derived mesenchymal stem cells because they represent the most accessible source of stem cells in the human mouth.

  18. Genetic engineering and sustainable production of ornamentals

    DEFF Research Database (Denmark)

    Lütken, Henrik Vlk; Clarke, Jihong Liu; Müller, Renate

    2012-01-01

    and reduction of chemicals applied during production of ornamental plants. Numerous chemicals used in modern plant production have negative impacts on human health and are hazardous to the environment. In Europe, several compounds have lost their approval and further legal restrictions can be expected....... This review presents the more recent progress of genetic engineering in ornamental breeding, delivers an overview of the biological background of the used technologies and critically evaluates the usefulness of the strategies to obtain improved ornamental plants. First, genetic engineering is addressed......Abstract Through the last decades, environmentally and health-friendly production methods and conscientious use of resources have become crucial for reaching the goal of a more sustainable plant production. Protection of the environment requires careful consumption of limited resources...

  19. Genetic engineering of cyanobacteria as biodiesel feedstock.

    Energy Technology Data Exchange (ETDEWEB)

    Ruffing, Anne.; Trahan, Christine Alexandra; Jones, Howland D. T.

    2013-01-01

    Algal biofuels are a renewable energy source with the potential to replace conventional petroleum-based fuels, while simultaneously reducing greenhouse gas emissions. The economic feasibility of commercial algal fuel production, however, is limited by low productivity of the natural algal strains. The project described in this SAND report addresses this low algal productivity by genetically engineering cyanobacteria (i.e. blue-green algae) to produce free fatty acids as fuel precursors. The engineered strains were characterized using Sandias unique imaging capabilities along with cutting-edge RNA-seq technology. These tools are applied to identify additional genetic targets for improving fuel production in cyanobacteria. This proof-of-concept study demonstrates successful fuel production from engineered cyanobacteria, identifies potential limitations, and investigates several strategies to overcome these limitations. This project was funded from FY10-FY13 through the President Harry S. Truman Fellowship in National Security Science and Engineering, a program sponsored by the LDRD office at Sandia National Laboratories.

  20. Genetic Engineering and Competitiveness of Livestock Production

    Directory of Open Access Journals (Sweden)

    Carl A.Pinkert

    2003-06-01

    Full Text Available Our ability to modify whole animal genetics has grown considerably in the last two decades. We have seen concerns regarding food safety and protection of breeding rights of genetically modified animals compel redirection of genetic engineering experimentation toward biomedical applications. Indeed, it has been nearly twenty years since the first transgenic livestock appeared in the literature, yet at this time, there are no commercially viable agricultural species. In contrast to commercialization concerns, in a variety of existing transgenic animal models, basic research into the regulation and function of specific genes (including both gain-of-function and ablation of potentially deleterious gene products has persevered. Pioneering efforts in transgenic animal technology have markedly influenced our appreciation of the factors that govern gene regulation and expression, and have contributed significantly to our understanding of the biology of mammalian development.

  1. Muscle Tissue Engineering Using Gingival Mesenchymal Stem Cells Encapsulated in Alginate Hydrogels Containing Multiple Growth Factors.

    Science.gov (United States)

    Ansari, Sahar; Chen, Chider; Xu, Xingtian; Annabi, Nasim; Zadeh, Homayoun H; Wu, Benjamin M; Khademhosseini, Ali; Shi, Songtao; Moshaverinia, Alireza

    2016-06-01

    Repair and regeneration of muscle tissue following traumatic injuries or muscle diseases often presents a challenging clinical situation. If a significant amount of tissue is lost the native regenerative potential of skeletal muscle will not be able to grow to fill the defect site completely. Dental-derived mesenchymal stem cells (MSCs) in combination with appropriate scaffold material, present an advantageous alternative therapeutic option for muscle tissue engineering in comparison to current treatment modalities available. To date, there has been no report on application of gingival mesenchymal stem cells (GMSCs) in three-dimensional scaffolds for muscle tissue engineering. The objectives of the current study were to develop an injectable 3D RGD-coupled alginate scaffold with multiple growth factor delivery capacity for encapsulating GMSCs, and to evaluate the capacity of encapsulated GMSCs to differentiate into myogenic tissue in vitro and in vivo where encapsulated GMSCs were transplanted subcutaneously into immunocompromised mice. The results demonstrate that after 4 weeks of differentiation in vitro, GMSCs as well as the positive control human bone marrow mesenchymal stem cells (hBMMSCs) exhibited muscle cell-like morphology with high levels of mRNA expression for gene markers related to muscle regeneration (MyoD, Myf5, and MyoG) via qPCR measurement. Our quantitative PCR analyzes revealed that the stiffness of the RGD-coupled alginate regulates the myogenic differentiation of encapsulated GMSCs. Histological and immunohistochemical/fluorescence staining for protein markers specific for myogenic tissue confirmed muscle regeneration in subcutaneous transplantation in our in vivo animal model. GMSCs showed significantly greater capacity for myogenic regeneration in comparison to hBMMSCs (p engineering.

  2. Sternal Repair with Bone Grafts Engineered from Amniotic Mesenchymal Stem Cells

    Science.gov (United States)

    Steigman, Shaun A.; Ahmed, Azra; Shanti, Rabie M.; Tuan, Rocky S.; Valim, Clarissa; Fauza, Dario O.

    2013-01-01

    Background We aimed at determining whether osseous grafts engineered from amniotic mesenchymal stem cells (aMSCs) could be employed in postnatal sternal repair. Methods Leporine aMSCs were isolated, identified, transfected with green fluorescent protein (GFP), expanded, and seeded onto biodegradable electrospun nanofibrous scaffolds (n=6). Constructs were dynamically maintained in an osteogenic medium and equally divided into two groups with respect to time in vitro, namely 14.6 or 33.9 weeks. They were then used to repair full thickness sternal defects spanning 2–3 intercostal spaces in allogeneic kits (n=6). Grafts were submitted to multiple analyses 2 months thereafter. Results Chest roentgenograms showed defect closure in all animals, confirmed at necropsy. Graft density as assessed by micro-CT scans increased significantly in vivo, yet there were no differences in mineralization by extracellular calcium measurements pre- and post-implantation. There was a borderline increase in alkaline phosphatase activity in vivo, suggesting ongoing graft remodeling. Histologically, implants contained GFP-positive cells and few mononuclear infiltrates. There were no differences between the two construct groups in any comparison. Conclusions Engineered osseous grafts derived from amniotic mesenchymal stem cells may become a viable alternative for sternal repair. The amniotic fluid can be a practical cell source for engineered chest wall reconstruction. PMID:19524727

  3. Genetic engineering of microorganisms for biodiesel production.

    Science.gov (United States)

    Lin, Hui; Wang, Qun; Shen, Qi; Zhan, Jumei; Zhao, Yuhua

    2013-01-01

    Biodiesel, as one type of renewable energy, is an ideal substitute for petroleum-based diesel fuel and is usually made from triacylglycerides by transesterification with alcohols. Biodiesel production based on microbial fermentation aiming to establish more efficient, less-cost and sustainable biodiesel production strategies is under current investigation by various start-up biotechnology companies and research centers. Genetic engineering plays a key role in the transformation of microbes into the desired cell factories with high efficiency of biodiesel production. Here, we present an overview of principal microorganisms used in the microbial biodiesel production and recent advances in metabolic engineering for the modification required. Overexpression or deletion of the related enzymes for de novo synthesis of biodiesel is highlighted with relevant examples.

  4. Advances in genetic engineering of marine algae.

    Science.gov (United States)

    Qin, Song; Lin, Hanzhi; Jiang, Peng

    2012-01-01

    Algae are a component of bait sources for animal aquaculture, and they produce abundant valuable compounds for the chemical industry and human health. With today's fast growing demand for algae biofuels and the profitable market for cosmetics and pharmaceuticals made from algal natural products, the genetic engineering of marine algae has been attracting increasing attention as a crucial systemic technology to address the challenge of the biomass feedstock supply for sustainable industrial applications and to modify the metabolic pathway for the more efficient production of high-value products. Nevertheless, to date, only a few marine algae species can be genetically manipulated. In this article, an updated account of the research progress in marine algal genomics is presented along with methods for transformation. In addition, vector construction and gene selection strategies are reviewed. Meanwhile, a review on the progress of bioreactor technologies for marine algae culture is also revisited. Copyright © 2012 Elsevier Inc. All rights reserved.

  5. Genetically engineering adenoviral vectors for gene therapy.

    Science.gov (United States)

    Coughlan, Lynda

    2014-01-01

    Adenoviral (Ad) vectors are commonly used for various gene therapy applications. Significant advances in the genetic engineering of Ad vectors in recent years has highlighted their potential for the treatment of metastatic disease. There are several methods to genetically modify the Ad genome to incorporate retargeting peptides which will redirect the natural tropism of the viruses, including homologous recombination in bacteria or yeast. However, homologous recombination in yeast is highly efficient and can be achieved without the need for extensive cloning strategies. In addition, the method does not rely on the presence of unique restriction sites within the Ad genome and the reagents required for this method are widely available and inexpensive. Large plasmids containing the entire adenoviral genome (~36 kbp) can be modified within Saccharomyces cerevisiae yeast and genomes easily rescued in Escherichia coli hosts for analysis or amplification. A method for two-step homologous recombination in yeast is described in this chapter.

  6. Xenomicrobiology: a roadmap for genetic code engineering.

    Science.gov (United States)

    Acevedo-Rocha, Carlos G; Budisa, Nediljko

    2016-09-01

    Biology is an analytical and informational science that is becoming increasingly dependent on chemical synthesis. One example is the high-throughput and low-cost synthesis of DNA, which is a foundation for the research field of synthetic biology (SB). The aim of SB is to provide biotechnological solutions to health, energy and environmental issues as well as unsustainable manufacturing processes in the frame of naturally existing chemical building blocks. Xenobiology (XB) goes a step further by implementing non-natural building blocks in living cells. In this context, genetic code engineering respectively enables the re-design of genes/genomes and proteins/proteomes with non-canonical nucleic (XNAs) and amino (ncAAs) acids. Besides studying information flow and evolutionary innovation in living systems, XB allows the development of new-to-nature therapeutic proteins/peptides, new biocatalysts for potential applications in synthetic organic chemistry and biocontainment strategies for enhanced biosafety. In this perspective, we provide a brief history and evolution of the genetic code in the context of XB. We then discuss the latest efforts and challenges ahead for engineering the genetic code with focus on substitutions and additions of ncAAs as well as standard amino acid reductions. Finally, we present a roadmap for the directed evolution of artificial microbes for emancipating rare sense codons that could be used to introduce novel building blocks. The development of such xenomicroorganisms endowed with a 'genetic firewall' will also allow to study and understand the relation between code evolution and horizontal gene transfer. © 2016 The Authors. Microbial Biotechnology published by John Wiley & Sons Ltd and Society for Applied Microbiology.

  7. Engineering tubular bone using mesenchymal stem cell sheets and coral particles

    Energy Technology Data Exchange (ETDEWEB)

    Geng, Wenxin [Key Laboratory of Resource Biology and Biotechnology in Western China, Ministry of Education, College of Life Science, Northwest University, No.229 North Taibai Road, Xi’an 710069 (China); Ma, Dongyang [Department of Oral and Maxillofacial Surgery, Lanzhou General Hospital, Lanzhou Command of PLA, BinHe 333 South Road, Lanzhou 730052 (China); Yan, Xingrong; Liu, Liangqi; Cui, Jihong; Xie, Xin; Li, Hongmin [Key Laboratory of Resource Biology and Biotechnology in Western China, Ministry of Education, College of Life Science, Northwest University, No.229 North Taibai Road, Xi’an 710069 (China); Chen, Fulin, E-mail: chenfl@nwu.edu.cn [Key Laboratory of Resource Biology and Biotechnology in Western China, Ministry of Education, College of Life Science, Northwest University, No.229 North Taibai Road, Xi’an 710069 (China)

    2013-04-19

    Highlights: • We developed a novel engineering strategy to solve the limitations of bone grafts. • We fabricated tubular constructs using cell sheets and coral particles. • The composite constructs showed high radiological density and compressive strength. • These characteristics were similar to those of native bone. -- Abstract: The development of bone tissue engineering has provided new solutions for bone defects. However, the cell-scaffold-based approaches currently in use have several limitations, including low cell seeding rates and poor bone formation capacity. In the present study, we developed a novel strategy to engineer bone grafts using mesenchymal stem cell sheets and coral particles. Rabbit bone marrow mesenchymal stem cells were continuously cultured to form a cell sheet with osteogenic potential and coral particles were integrated into the sheet. The composite sheet was then wrapped around a cylindrical mandrel to fabricate a tubular construct. The resultant tubular construct was cultured in a spinner-flask bioreactor and subsequently implanted into a subcutaneous pocket in a nude mouse for assessment of its histological characteristics, radiological density and mechanical property. A similar construct assembled from a cell sheet alone acted as a control. In vitro observations demonstrated that the composite construct maintained its tubular shape, and exhibited higher radiological density, compressive strength and greater extracellular matrix deposition than did the control construct. In vivo experiments further revealed that new bone formed ectopically on the composite constructs, so that the 8-week explants of the composite sheets displayed radiological density similar to that of native bone. These results indicate that the strategy of using a combination of a cell sheet and coral particles has great potential for bone tissue engineering and repairing bone defects.

  8. Can Man Control His Biological Evolution? A Symposium on Genetic Engineering. Genetic Engineering

    Science.gov (United States)

    Ramsey, Paul

    1972-01-01

    Presented are issues related to genetic engineering. Increased knowledge of techniques to manipulate genes are apt to create confusion about moral values in relation to unborn babies and other living organisms on earth. Human beings may use this knowledge to disturb the balance maintained by nature. (PS)

  9. Effects of mechanical loading on human mesenchymal stem cells for cartilage tissue engineering.

    Science.gov (United States)

    Choi, Jane Ru; Yong, Kar Wey; Choi, Jean Yu

    2017-05-19

    Today, articular cartilage damage is a major health problem, affecting people of all ages. The existing conventional articular cartilage repair techniques, such as autologous chondrocyte implantation (ACI), microfracture, and mosaicplasty, have many shortcomings which negatively affect their clinical outcomes. Therefore, it is essential to develop an alternative and efficient articular repair technique that can address those shortcomings. Cartilage tissue engineering, which aims to create a tissue-engineered cartilage derived from human mesenchymal stem cells (MSCs), shows great promise for improving articular cartilage defect therapy. However, the use of tissue-engineered cartilage for the clinical therapy of articular cartilage defect still remains challenging. Despite the importance of mechanical loading to create a functional cartilage has been well demonstrated, the specific type of mechanical loading and its optimal loading regime is still under investigation. This review summarizes the most recent advances in the effects of mechanical loading on human MSCs. First, the existing conventional articular repair techniques and their shortcomings are highlighted. The important parameters for the evaluation of the tissue-engineered cartilage, including chondrogenic and hypertrophic differentiation of human MSCs are briefly discussed. The influence of mechanical loading on human MSCs is subsequently reviewed and the possible mechanotransduction signaling is highlighted. The development of non-hypertrophic chondrogenesis in response to the changing mechanical microenvironment will aid in the establishment of a tissue-engineered cartilage for efficient articular cartilage repair. © 2017 Wiley Periodicals, Inc.

  10. Genetic engineering with T cell receptors.

    Science.gov (United States)

    Zhang, Ling; Morgan, Richard A

    2012-06-01

    In the past two decades, human gene transfer research has been translated from a laboratory technology to clinical evaluation. The success of adoptive transfer of tumor-reactive lymphocytes to treat the patients with metastatic melanoma has led to new strategies to redirect normal T cells to recognize tumor antigens by genetic engineering with tumor antigen-specific T cell receptor (TCR) genes. This new strategy can generate large numbers of defined antigen-specific cells for therapeutic application. Much progress has been made to TCR gene transfer systems by optimizing gene expression and gene transfer protocols. Vector and protein modifications have enabled excellent expression of introduced TCR chains in human lymphocytes with reduced mis-pairing between the introduced and endogenous TCR chains. Initial clinical studies have demonstrated that TCR gene-engineered T cells could mediate tumor regression in vivo. In this review, we discuss the progress and prospects of TCR gene-engineered T cells as a therapeutic strategy for treating patients with melanoma and other cancers.

  11. Human Genetic Engineering: A Survey of Student Value Stances

    Science.gov (United States)

    Wilson, Sara McCormack; And Others

    1975-01-01

    Assesses the values of high school and college students relative to human genetic engineering and recommends that biology educators explore instructional strategies merging human genetic information with value clarification techniques. (LS)

  12. Seeking perfection: a Kantian look at human genetic engineering.

    Science.gov (United States)

    Gunderson, Martin

    2007-01-01

    It is tempting to argue that Kantian moral philosophy justifies prohibiting both human germ-line genetic engineering and non-therapeutic genetic engineering because they fail to respect human dignity. There are, however, good reasons for resisting this temptation. In fact, Kant's moral philosophy provides reasons that support genetic engineering-even germ-line and non-therapeutic. This is true of Kant's imperfect duties to seek one's own perfection and the happiness of others. It is also true of the categorical imperative. Kant's moral philosophy does, however, provide limits to justifiable genetic engineering.

  13. The efficacy of polycaprolactone/hydroxyapatite scaffold in combination with mesenchymal stem cells for bone tissue engineering.

    Science.gov (United States)

    Chuenjitkuntaworn, Boontharika; Osathanon, Thanaphum; Nowwarote, Nunthawan; Supaphol, Pitt; Pavasant, Prasit

    2016-01-01

    Major drawbacks of using an autograft are the possibilities of insufficient bony source and patient's morbidity after operation. Bone tissue engineering technology, therefore, has been applied for repairing bony defects. Previous study showed that a novel fabricated 3D-Polycaprolactone/Hydroxyapatite (PCL/HAp) scaffold possessed a good biocompatibility for bone cells. This study aimed to determine the ability of PCL/HAp for supporting cell growth, gene expression, and osteogenic differentiation in three types of mesenchymal stem cells, including bone marrow-derived mesenchymal stem cells (BMSCs), dental pulp stem cells (DPSCs), and adiposed-derived mesenchymal stem cells (ADSCs). These were assessed by cell viability assay (MTT), reverse-transcription polymerase chain reaction (RT-PCR) analysis, alkaline phosphatase activity, and osteogenic differentiation by alizarin red-S staining. The results showed that PCL/HAp scaffold could support growth of all three types of mesenchymal stem cells. In addition, DPSCs with PCL/HAp showed the highest level of calcium deposition compared to other groups. In conclusion, DPSCs exhibited a better compatibility with these scaffolds compared to BMSCs and ADSCs. However, the PCL/HAp could be a good candidate scaffold for all tested mesenchymal stem cells in bone tissue engineering. © 2015 Wiley Periodicals, Inc.

  14. Improved Quantum Genetic Algorithm in Application of Scheduling Engineering Personnel

    Directory of Open Access Journals (Sweden)

    Huaixiao Wang

    2014-01-01

    Full Text Available To verify the availability of the improved quantum genetic algorithm in solving the scheduling engineering personnel problem, the following work has been carried out: the characteristics of the scheduling engineering personnel problem are analyzed, the quantum encoding method is proposed, and an improved quantum genetic algorithm is applied to address the issue. Taking the low efficiency and the bad performance of the conventional quantum genetic algorithm into account, a universal improved quantum genetic algorithm is introduced to solve the scheduling engineering personnel problem. Finally, the examples are applied to verify the effectiveness and superiority of the improved quantum genetic algorithm and the rationality of the encoding method.

  15. Programmable genetic circuits for pathway engineering.

    Science.gov (United States)

    Hoynes-O'Connor, Allison; Moon, Tae Seok

    2015-12-01

    Synthetic biology has the potential to provide decisive advances in genetic control of metabolic pathways. However, there are several challenges that synthetic biologists must overcome before this vision becomes a reality. First, a library of diverse and well-characterized sensors, such as metabolite-sensing or condition-sensing promoters, must be constructed. Second, robust programmable circuits that link input conditions with a specific gene regulation response must be developed. Finally, multi-gene targeting strategies must be integrated with metabolically relevant sensors and complex, robust logic. Achievements in each of these areas, which employ the CRISPR/Cas system, in silico modeling, and dynamic sensor-regulators, among other tools, provide a strong basis for future research. Overall, the future for synthetic biology approaches in metabolic engineering holds immense promise.

  16. Modularization of genetic elements promotes synthetic metabolic engineering.

    Science.gov (United States)

    Qi, Hao; Li, Bing-Zhi; Zhang, Wen-Qian; Liu, Duo; Yuan, Ying-Jin

    2015-11-15

    In the context of emerging synthetic biology, metabolic engineering is moving to the next stage powered by new technologies. Systematical modularization of genetic elements makes it more convenient to engineer biological systems for chemical production or other desired purposes. In the past few years, progresses were made in engineering metabolic pathway using synthetic biology tools. Here, we spotlighted the topic of implementation of modularized genetic elements in metabolic engineering. First, we overviewed the principle developed for modularizing genetic elements and then discussed how the genetic modules advanced metabolic engineering studies. Next, we picked up some milestones of engineered metabolic pathway achieved in the past few years. Last, we discussed the rapid raised synthetic biology field of "building a genome" and the potential in metabolic engineering.

  17. Cartilage tissue engineering: Role of mesenchymal stem cells along with growth factors & scaffolds

    Directory of Open Access Journals (Sweden)

    M B Gugjoo

    2016-01-01

    Full Text Available Articular cartilage injury poses a major challenge for both the patient and orthopaedician. Articular cartilage defects once formed do not regenerate spontaneously, rather replaced by fibrocartilage which is weaker in mechanical competence than the normal hyaline cartilage. Mesenchymal stem cells (MSCs along with different growth factors and scaffolds are currently incorporated in tissue engineering to overcome the deficiencies associated with currently available surgical methods and to facilitate cartilage healing. MSCs, being readily available with a potential to differentiate into chondrocytes which are enhanced by the application of different growth factors, are considered for effective repair of articular cartilage after injury. However, therapeutic application of MSCs and growth factors for cartilage repair remains in its infancy, with no comparative clinical study to that of the other surgical techniques. The present review covers the role of MSCs, growth factors and scaffolds for the repair of articular cartilage injury.

  18. Biodegradable Thermogel as Culture Matrix of Bone Marrow Mesenchymal Stem Cells for Potential Cartilage Tissue Engineering

    Institute of Scientific and Technical Information of China (English)

    Yan-bo Zhang; Jian-xun Ding; Wei-guo Xu; Jie Wu; Fei Chang; Xiu-li Zhuang; Xue-si Chen

    2014-01-01

    Poly(lactide-co-glycolide)-poly(ethylene glycol)-poly(lactide-co-glycolide) (PLGA-PEG-PLGA) triblock copolymer was synthesized through the ring-opening polymerization of LA and GA with PEG as macroinitiator and stannous octoate as catalyst.The amphiphilic copolymer self-assembled into micelles in aqueous solutions,and formed hydrogels as the increase of temperature at relatively high concentrations (> 15 wt%).The favorable degradability of the hydrogel was confirmed by in vitro and in vivo degradation experiments.The good cellular and tissular compatibilities of the thermogel were demonstrated.The excellent adhesion and proliferation of bone marrow mesenchymal stem cells endowed PLGA-PEG-PLGA thermogelling hydrogel with fascinating prospect for cartilage tissue engineering.

  19. Antibody-engineered nanoparticles selectively inhibit mesenchymal cells isolated from patients with chronic lung allograft dysfunction.

    Science.gov (United States)

    Cova, Emanuela; Colombo, Miriam; Inghilleri, Simona; Morosini, Monica; Miserere, Simona; Peñaranda-Avila, Jesus; Santini, Benedetta; Piloni, Davide; Magni, Sara; Gramatica, Furio; Prosperi, Davide; Meloni, Federica

    2015-01-01

    Chronic lung allograft dysfunction represents the main cause of death after lung transplantation, and so far there is no effective therapy. Mesenchymal cells (MCs) are primarily responsible for fibrous obliteration of small airways typical of chronic lung allograft dysfunction. Here, we engineered gold nanoparticles containing a drug in the hydrophobic section to inhibit MCs, and exposing on the outer hydrophilic surface a monoclonal antibody targeting a MC-specific marker (half-chain gold nanoparticles with everolimus). Half-chain gold nanoparticles with everolimus have been synthesized and incubated with MCs to evaluate the effect on proliferation and apoptosis. Drug-loaded gold nanoparticles coated with the specific antibody were able to inhibit proliferation and induce apoptosis without stimulating an inflammatory response, as assessed by in vitro experiments. These findings demonstrate the effectiveness of our nanoparticles in inhibiting MCs and open new perspectives for a local treatment of chronic lung allograft dysfunction.

  20. PUBLIC PERCEPTION OF GENETIC ENGINEERING AND THE CHOICE TO PURCHASE GENETICALLY MODIFIED FOOD

    OpenAIRE

    2004-01-01

    This paper presents the results of a survey conducted on public perception of genetic engineering in Jamaica. Our findings suggest that the safety of genetically modified foods is a major concern for consumers and that the perception of the prospects for genetic engineering to improve the quality of life represents a major factor in a consumer's decision to purchase GM foods.

  1. Use of Genetically Modified Mesenchymal Stem Cells to Treat Neurodegenerative Diseases

    Directory of Open Access Journals (Sweden)

    Robert D. Wyse

    2014-01-01

    Full Text Available The transplantation of mesenchymal stem cells (MSCs for treating neurodegenerative disorders has received growing attention recently because these cells are readily available, easily expanded in culture, and when transplanted, survive for relatively long periods of time. Given that such transplants have been shown to be safe in a variety of applications, in addition to recent findings that MSCs have useful immunomodulatory and chemotactic properties, the use of these cells as vehicles for delivering or producing beneficial proteins for therapeutic purposes has been the focus of several labs. In our lab, the use of genetic modified MSCs to release neurotrophic factors for the treatment of neurodegenerative diseases is of particular interest. Specifically, glial cell-derived neurotrophic factor (GDNF, nerve growth factor (NGF, and brain derived neurotrophic factor (BDNF have been recognized as therapeutic trophic factors for Parkinson’s, Alzheimer’s and Huntington’s diseases, respectively. The aim of this literature review is to provide insights into: (1 the inherent properties of MSCs as a platform for neurotrophic factor delivery; (2 the molecular tools available for genetic manipulation of MSCs; (3 the rationale for utilizing various neurotrophic factors for particular neurodegenerative diseases; and (4 the clinical challenges of utilizing genetically modified MSCs.

  2. Use of genetically modified mesenchymal stem cells to treat neurodegenerative diseases.

    Science.gov (United States)

    Wyse, Robert D; Dunbar, Gary L; Rossignol, Julien

    2014-01-23

    The transplantation of mesenchymal stem cells (MSCs) for treating neurodegenerative disorders has received growing attention recently because these cells are readily available, easily expanded in culture, and when transplanted, survive for relatively long periods of time. Given that such transplants have been shown to be safe in a variety of applications, in addition to recent findings that MSCs have useful immunomodulatory and chemotactic properties, the use of these cells as vehicles for delivering or producing beneficial proteins for therapeutic purposes has been the focus of several labs. In our lab, the use of genetic modified MSCs to release neurotrophic factors for the treatment of neurodegenerative diseases is of particular interest. Specifically, glial cell-derived neurotrophic factor (GDNF), nerve growth factor (NGF), and brain derived neurotrophic factor (BDNF) have been recognized as therapeutic trophic factors for Parkinson's, Alzheimer's and Huntington's diseases, respectively. The aim of this literature review is to provide insights into: (1) the inherent properties of MSCs as a platform for neurotrophic factor delivery; (2) the molecular tools available for genetic manipulation of MSCs; (3) the rationale for utilizing various neurotrophic factors for particular neurodegenerative diseases; and (4) the clinical challenges of utilizing genetically modified MSCs.

  3. Plant Genetic Resources: Selected Issues from Genetic Erosion to Genetic Engineering

    Directory of Open Access Journals (Sweden)

    Karl Hammer

    2008-04-01

    Full Text Available Plant Genetic Resources (PGR continue to play an important role in the development of agriculture. The following aspects receive a special consideration:1. Definition. The term was coined in 1970. The genepool concept served as an important tool in the further development. Different approaches are discussed.2. Values of Genetic Resources. A short introduction is highlighting this problem and stressing the economic usfulness of PGR.3. Genetic Erosion. Already observed by E. Baur in 1914, this is now a key issue within PGR. The case studies cited include Ethiopia, Italy, China, S Korea, Greece and S. Africa. Modern approaches concentrate on allelic changes in varieties over time but neglect the landraces. The causes and consequences of genetic erosion are discussed.4. Genetic Resources Conservation. Because of genetic erosion there is a need for conservation. PGR should be consigned to the appropriate method of conservation (ex situ, in situ, on-farm according to the scientific basis of biodiversity (genetic diversity, species diversity, ecosystem diversity and the evolutionary status of plants (cultivated plants, weeds, related wild plants (crop wild relatives.5. GMO. The impact of genetically engineered plants on genetic diversity is discussed.6. The Conclusions and Recommendations stress the importance of PGR. Their conservation and use are urgent necessities for the present development and future survival of mankind.

  4. Improved Quantum Genetic Algorithm in Application of Scheduling Engineering Personnel

    OpenAIRE

    Huaixiao Wang; Ling Li; Jianyong Liu; Yong Wang; Chengqun Fu

    2014-01-01

    To verify the availability of the improved quantum genetic algorithm in solving the scheduling engineering personnel problem, the following work has been carried out: the characteristics of the scheduling engineering personnel problem are analyzed, the quantum encoding method is proposed, and an improved quantum genetic algorithm is applied to address the issue. Taking the low efficiency and the bad performance of the conventional quantum genetic algorithm into account, a universal improved q...

  5. Engineered Plant Minichromosome and Its Application in Genomics and Genetic Engineering

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    Engineered minichromosomes have been constructed as novel artificial chromosome platforms for future genetic engineering in maize.We demonstrated that minichromosomes could be created by telomere-mediated chromosomal truncation of both normal A chromosomes and the supernumerary B

  6. Efforts and Challenges in Engineering the Genetic Code.

    Science.gov (United States)

    Lin, Xiao; Yu, Allen Chi Shing; Chan, Ting Fung

    2017-03-14

    This year marks the 48th anniversary of Francis Crick's seminal work on the origin of the genetic code, in which he first proposed the "frozen accident" hypothesis to describe evolutionary selection against changes to the genetic code that cause devastating global proteome modification. However, numerous efforts have demonstrated the viability of both natural and artificial genetic code variations. Recent advances in genetic engineering allow the creation of synthetic organisms that incorporate noncanonical, or even unnatural, amino acids into the proteome. Currently, successful genetic code engineering is mainly achieved by creating orthogonal aminoacyl-tRNA/synthetase pairs to repurpose stop and rare codons or to induce quadruplet codons. In this review, we summarize the current progress in genetic code engineering and discuss the challenges, current understanding, and future perspectives regarding genetic code modification.

  7. Efforts and Challenges in Engineering the Genetic Code

    Directory of Open Access Journals (Sweden)

    Xiao Lin

    2017-03-01

    Full Text Available This year marks the 48th anniversary of Francis Crick’s seminal work on the origin of the genetic code, in which he first proposed the “frozen accident” hypothesis to describe evolutionary selection against changes to the genetic code that cause devastating global proteome modification. However, numerous efforts have demonstrated the viability of both natural and artificial genetic code variations. Recent advances in genetic engineering allow the creation of synthetic organisms that incorporate noncanonical, or even unnatural, amino acids into the proteome. Currently, successful genetic code engineering is mainly achieved by creating orthogonal aminoacyl-tRNA/synthetase pairs to repurpose stop and rare codons or to induce quadruplet codons. In this review, we summarize the current progress in genetic code engineering and discuss the challenges, current understanding, and future perspectives regarding genetic code modification.

  8. Scaffold-free parathyroid tissue engineering using tonsil-derived mesenchymal stem cells.

    Science.gov (United States)

    Park, Yoon Shin; Hwang, Ji-Young; Jun, Yesl; Jin, Yoon Mi; Kim, Gyungah; Kim, Ha Yeong; Kim, Han Su; Lee, Sang-Hoon; Jo, Inho

    2016-04-15

    To restore damaged parathyroid function, parathyroid tissue engineering is the best option. Previously, we reported that differentiated tonsil-derived mesenchymal stem cells (dTMSC) restore in vivo parathyroid function, but only if they are embedded in a scaffold. Because of the limited biocompatibility of Matrigel, however, here we developed a more clinically applicable, scaffold-free parathyroid regeneration system. Scaffold-free dTMSC spheroids were engineered in concave microwell plates made of polydimethylsiloxane in control culture medium for the first 7days and differentiation medium (containing activin A and sonic hedgehog) for next 7days. The size of dTMSC spheroids showed a gradual and significant decrease up to day 5, whereafter it decreased much less. Cells in dTMSC spheroids were highly viable (>80%). They expressed high levels of intact parathyroid hormone (iPTH), the parathyroid secretory protein 1, and cell adhesion molecule, N-cadherin. Furthermore, dTMSC spheroids-implanted parathyroidectomized (PTX) rats revealed higher survival rates (50%) over a 3-month period with physiological levels of both serum iPTH (57.7-128.2pg/mL) and ionized calcium (0.70-1.15mmol/L), compared with PTX rats treated with either vehicle or undifferentiated TMSC spheroids. This is the first report of a scaffold-free, human stem cell-based parathyroid tissue engineering and represents a more clinically feasible strategy for hypoparathyroidism treatment than those requiring scaffolds. Herein, we have for the first time developed a scaffold-free parathyroid tissue spheroids using differentiated tonsil-derived mesenchymal stem cells (dTMSC) to restore in vivo parathyroid cell functions. This new strategy is effective, even for long periods (3months), and is thus likely to be more feasible in clinic for hypoparathyroidism treatment. Development of TMSC spheroids may also provide a convenient and efficient scaffold-free platform for researchers investigating conditions

  9. Engineering anisotropic biomimetic fibrocartilage microenvironment by bioprinting mesenchymal stem cells in nanoliter gel droplets.

    Science.gov (United States)

    Gurkan, Umut A; El Assal, Rami; Yildiz, Simin E; Sung, Yuree; Trachtenberg, Alexander J; Kuo, Winston P; Demirci, Utkan

    2014-07-07

    Over the past decade, bioprinting has emerged as a promising patterning strategy to organize cells and extracellular components both in two and three dimensions (2D and 3D) to engineer functional tissue mimicking constructs. So far, tissue printing has neither been used for 3D patterning of mesenchymal stem cells (MSCs) in multiphase growth factor embedded 3D hydrogels nor been investigated phenotypically in terms of simultaneous differentiation into different cell types within the same micropatterned 3D tissue constructs. Accordingly, we demonstrated a biochemical gradient by bioprinting nanoliter droplets encapsulating human MSCs, bone morphogenetic protein 2 (BMP-2), and transforming growth factor β1 (TGF- β1), engineering an anisotropic biomimetic fibrocartilage microenvironment. Assessment of the model tissue construct displayed multiphasic anisotropy of the incorporated biochemical factors after patterning. Quantitative real time polymerase chain reaction (qRT-PCR) results suggested genomic expression patterns leading to simultaneous differentiation of MSC populations into osteogenic and chondrogenic phenotype within the multiphasic construct, evidenced by upregulation of osteogenesis and condrogenesis related genes during in vitro culture. Comprehensive phenotypic network and pathway analysis results, which were based on genomic expression data, indicated activation of differentiation related mechanisms, via signaling pathways, including TGF, BMP, and vascular endothelial growth factor.

  10. Full-thickness tissue engineered skin constructed with autogenic bone marrow mesenchymal stem cells

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    To explore the feasibility of repairing clinical cutaneous deficiency, autogenic bone marrow mesen-chymal stem cells (BMSCs) were isolated and differentiated into epidermal cells and fibroblasts in vitro supplemented with different inducing factors and biomaterials to construct functional tissue- engineered skin. The results showed that after 72 h induction, BMSCs displayed morphologic changes such as typical epidermal cell arrangement, from spindle shape to round or oval; tonofibrils, melano-somes and keratohyaline granules were observed under a transmission electronic microscope. The differentiated cells expressed epidermal stem cell surface marker CK19 (59.66% ± 4.2%) and epidermal cells differentiation marker CK10. In addition, the induced epidermal cells acquired the anti-radiation capacity featured by lowered apoptosis following exposure to UVB. On the other hand, the collagen microfibrils deposition was noticed under a transmission electronic microscope after differentiating into dermis fibroblasts; RT-PCR identified collagen type I mRNA expression in differentiated cells; radioimmunoassay detected the secretion of interleukin-6 (IL-6) and interleukin-8 (IL-8) (up to 115.06 pg/mL and 0.84 ng/mL, respectively). Further in vivo implanting BMSCs with scaffold material short-ened skin wound repair significantly. In one word, autogenic BMSCs have the potential to differentiate into epidermal cells and fibroblasts in vitro, and show clinical feasibility acting as epidermis-like and dermis-like seed cells in skin engineering.

  11. Tissue engineering of rat bladder using marrow-derived mesenchymal stem cells and bladder acellular matrix.

    Directory of Open Access Journals (Sweden)

    Daniel L Coutu

    Full Text Available Bladder replacement or augmentation is required in congenital malformations or following trauma or cancer. The current surgical solution involves enterocystoplasty but is associated with high complication rates. Strategies for bladder tissue engineering are thus actively sought to address this unmet clinical need. Because of the poor efficacy of synthetic polymers, the use of bladder acellular matrix (BAM has been proposed. Indeed when cellular components are removed from xenogenic or allogeneic bladders, the extracellular matrix scaffold thus obtained can be used alone or in combination with stem cells. In this study, we propose the use of BAM seeded with marrow-derived mesenchymal stem cells (MSCs for bladder tissue engineering. We optimized a protocol for decellularization of bladder tissue from different species including rat, rabbit and swine. We demonstrate the use of non-ionic detergents followed by nuclease digestion results in efficient decellularization while preserving the extracellular matrix. When MSCs were seeded on acellular matrix scaffold, they remained viable and proliferative while adopting a cellular phenotype consistent with their microenvironment. Upon transplantation in rats after partial cystectomy, MSC-seeded BAM proved superior to unseeded BAM with animals recovering nearly 100% normal bladder capacity for up to six months. Histological analyses also demonstrated increased muscle regeneration.

  12. Spatial Engineering of Osteochondral Tissue Constructs Through Microfluidically Directed Differentiation of Mesenchymal Stem Cells.

    Science.gov (United States)

    Goldman, Stephen M; Barabino, Gilda A

    2016-01-01

    The development of tissue engineered osteochondral units has been slowed by a number of technical hurdles associated with recapitulating their heterogeneous nature ex vivo. Subsequently, numerous approaches with respect to cell sourcing, scaffolding composition, and culture media formulation have been pursued, which have led to high variability in outcomes and ultimately the lack of a consensus bioprocessing strategy. As such, the objective of this study was to standardize the design process by focusing on differentially supporting formation of cartilaginous and bony matrix by a single cell source in a spatially controlled manner within a single material system. A cell-polymer solution of bovine mesenchymal stem cells and agarose was cast against micromolds of a serpentine network and stacked to produce tissue constructs containing two independent microfluidic networks. Constructs were fluidically connected to two controlled flow loops and supplied with independently tuned differentiation parameters for chondrogenic and osteogenic induction, respectively. Constructs receiving inductive media showed differential gene expression of both chondrogenic and osteogenic markers in opposite directions along the thickness of the construct that was recapitulated at the protein level with respect to collagens I, II, and X. A control group receiving noninductive media showed homogeneous expression of these biomarkers measured in lower concentrations at both the mRNA and protein level. This work represents an important step in the rational design of engineered osteochondral units through establishment of an enabling technology for further optimization of scaffolding formulations and bioprocessing conditions toward the production of commercially viable osteochondral tissue products.

  13. Clinical Application of Human Mesenchymal Stromal Cells for Bone Tissue Engineering

    Directory of Open Access Journals (Sweden)

    Anindita Chatterjea

    2010-01-01

    Full Text Available The gold standard in the repair of bony defects is autologous bone grafting, even though it has drawbacks in terms of availability and morbidity at the harvesting site. Bone-tissue engineering, in which osteogenic cells and scaffolds are combined, is considered as a potential bone graft substitute strategy. Proof-of-principle for bone tissue engineering using mesenchymal stromal cells (MSCs has been demonstrated in various animal models. In addition, 7 human clinical studies have so far been conducted. Because the experimental design and evaluation parameters of the studies are rather heterogeneous, it is difficult to draw conclusive evidence on the performance of one approach over the other. However, it seems that bone apposition by the grafted MSCs in these studies is observed but not sufficient to bridge large bone defects. In this paper, we discuss the published human clinical studies performed so far for bone-tissue regeneration, using culture-expanded, nongenetically modified MSCs from various sources and extract from it points of consideration for future clinical studies.

  14. Scheduling in a Meta Search Engine by Genetic Algorithm

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    The meta search engines provide service to the users bydispensing the users' requests to the existing search engines. The existing search engines sele cted by meta search engine determine the searching quality. Because the performa nce of the existing search engines and the users' requests are changed dynamical ly, it is not favorable for the fixed search engines to optimize the holistic pe rformance of the meta search engine. This paper applies the genetic algorithm (G A) to realize the scheduling strategy of agent manager in our meta search engine , GSE(general search engine), which can simulate the evolution process of living things more lively and more efficiently. By using GA, the combination of search engines can be optimized and hence the holistic performance of GSE can be impro ved dramatically.

  15. Differentiation of mesenchymal stem cells into neuronal cells on fetal bovine acellular dermal matrix as a tissue engineered nerve scaffold

    Institute of Scientific and Technical Information of China (English)

    Yuping Feng; Jiao Wang; Shixin Ling; Zhuo Li; Mingsheng Li; Qiongyi Li; Zongren Ma; Sijiu Yu

    2014-01-01

    The purpose of this study was to assess fetal bovine acellular dermal matrix as a scaffold for supporting the differentiation of bone marrow mesenchymal stem cells into neural cells fol-lowing induction with neural differentiation medium. We performed long-term, continuous observation of cell morphology, growth, differentiation, and neuronal development using several microscopy techniques in conjunction with immunohistochemistry. We examined speciifc neu-ronal proteins and Nissl bodies involved in the differentiation process in order to determine the neuronal differentiation of bone marrow mesenchymal stem cells. The results show that bone marrow mesenchymal stem cells that differentiate on fetal bovine acellular dermal matrix display neuronal morphology with unipolar and bi/multipolar neurite elongations that express neuro-nal-speciifc proteins, includingβIII tubulin. The bone marrow mesenchymal stem cells grown on fetal bovine acellular dermal matrix and induced for long periods of time with neural differen-tiation medium differentiated into a multilayered neural network-like structure with long nerve ifbers that was composed of several parallel microifbers and neuronal cells, forming a complete neural circuit with dendrite-dendrite to axon-dendrite to dendrite-axon synapses. In addition, growth cones with filopodia were observed using scanning electron microscopy. Paraffin sec-tioning showed differentiated bone marrow mesenchymal stem cells with the typical features of neuronal phenotype, such as a large, round nucleus and a cytoplasm full of Nissl bodies. The data suggest that the biological scaffold fetal bovine acellular dermal matrix is capable of supporting human bone marrow mesenchymal stem cell differentiation into functional neurons and the subsequent formation of tissue engineered nerve.

  16. 骨髓间充质干细胞在骨组织工程中的应用%Bone marrow mesenchymal stem cell in bone tissue engineering

    Institute of Scientific and Technical Information of China (English)

    涂强; 徐国洲; 钟润泉; 王少华

    2006-01-01

    and culture of mesenchymal stem cells: density gradient centrifugation, flow cytometry and adherent screening method. Other scholars obtained cell strain of mesenchymal stem cell through single-cell cloning technique, but this way was more difficult to obtain multiple seed cells. At present, density gradient centrifugation was widely used. ② Mesenchymal stem cell could differentiate into bone cells and cartilae cells through adding inductor into eligible culture medium. Ideal scaffolds could carry and retain the vitality of cells, support rapid endogeny of vessels and be benefit for observing new bone formation under X ray. New bone might be absorbed and replaced in bone rebuilding and form or increase conductive bridging of host bone. Functions of differentiated cells could be maintained by the interaction between scaffolds surface and cells, and the histocompatibility was well. ③ Mesenchymal stem cell was an kind of ideal target cells for gene therapy. It could shift growth factor or cytokine and express foreign protein after multiple desintegration in vitro. It was successful in bone tissue engineering.CONCLUSION: Modern tissue engineering which is presented by stem cells develops quickly at present, but tissue engineering of mesenchymal stem cell starts just now. Bone marrow mesenchymal stem cell is characterized by easily materializing, potentiality of multiple tissue differentiation, stable genetic background, non-rejection in implant button and high proliferation. All properties determine that it will be a useful thing in cell and gene therapy as well as tissue engineering.

  17. NUTRITIONAL ENHANCEMENT OF ALFALFA THROUGH GENETIC ENGINEERING

    Directory of Open Access Journals (Sweden)

    J. Faragó

    2008-09-01

    Full Text Available Alfalfa (Medicago sativa L. is a pasture legume crop of primary importance to animal production throughout the world. The nutritional quality of alfalfa, as of other leguminous forage crops, is mainly determined by their content in selected essential amino acids (EAAs, such as methionine (Met and cysteine (Cys. In alfalfa, however, these S-containing amino acids constitute only about 1% or less of crude proteins (Frame et al., 1998. This is significantly less than the 3.5% Met+Cys content in the recommended FAO reference protein (FAO, 1973. Recent advances in genetic engineering allow to use the transgenic approach to increase the content of specific essential amino acids in target plant species. A number of different molecular approaches have been developed to address this issue, such as over-expression of a heterologous or homologous Met-rich protein, expression of a synthetic protein, modification of protein sequence, and metabolic engineering of the free amino acid pool and protein sink. To study the possibility of transgenic enhancement of nutritional quality of alfalfa, we used the approach of expression of a heterologous protein rich in Met+Cys in cells of alfalfa. The T-DNA introduced into the genome of alfalfa, using Agrobacterium tumefaciens-mediated genetic transformation, contained the selectable merker gene nptII for kanamycin (Kn resistance, and a cDNA of Ov gene from Japanese quail (Coturnix coturnix coding for a high Met+Cys containing ovalbumine (Mucha et al., 1991, both under constitutive promoters. After cocultivation of petiole segment- and leaf blade-explants of two highly embryogenic alfalfa genotypes Rg9/I-14-22 and Rg11/I-10-68 (Faragó et al., 1997 with cells of A. tumefaciens strain AGL1 carrying the nptII and Ov genes, and selection of transgenic cells on Kn containing selective media, more than one hundred putatively transgenic regenerants were obtained through somatic embryogenesis. Biological (Kn rooting assay

  18. Legal and regulatory aspects of genetically engineered animals.

    Science.gov (United States)

    Jones, D D

    1986-01-01

    The commercialization of genetically engineered food animals will pose a number of legal and regulatory questions. These may be grouped into questions of process and questions of products. The process of animal genetic engineering with artificially constructed vectors will probably be regulated in much the same manner as other veterinary procedures. There may be some discussion, however, as to whether animal drug or animal biologic regulations are more applicable. The products of animal genetic engineering, i.e., transgenic food animals and food products made from them, also raise important questions about product safety and identity. These include whether and how genetically engineered food animals will be subject to federal inspection for wholesomeness, whether artificial vectors, foreign genes, or gene products will adulterate recipient animal tissues, and how food products made from such animals will be labeled. Prior federal experience with the inspection of interspecific hybrids of cattle and buffalo provides a useful basis for further policy developments in the inspection and labeling of genetically engineered food animals. In particular, the inspection of cattle/buffalo hybrids has established a phenotypic (based on appearance) criterion for deciding how novel food animals should be inspected. As the genetic engineering of food animals on a production basis draws nearer, it may be necessary to supplement the phenotypic criterion with genetic (based on pedigree) criteria to assure that the essential characteristics of animals slaughtered under current food statutes are maintained.

  19. "Genetic Engineering" Gains Momentum (Science/Society Case Study).

    Science.gov (United States)

    Moore, John W.; Moore, Elizabeth A., Eds.

    1980-01-01

    Reviews the benefits and hazards of genetic engineering, or "recombinant-DNA" research. Recent federal safety rules issued by NIH which ease the strict prohibitions on recombinant-DNA research are explained. (CS)

  20. International Genetically Engineered Machine (iGEM) Competition

    CSIR Research Space (South Africa)

    Sparrow, RW

    2010-07-01

    Full Text Available iGEM, the International Genetically Engineered Machine competition, is an initiative from MIT and has become the premiere undergraduate synthetic biology competition. The competing teams consist of students who work on a synthetic biology project...

  1. Perspectives in Engineered Mesenchymal Stem/Stromal Cells Based Anti- Cancer Drug Delivery Systems.

    Science.gov (United States)

    Ackova, Darinka Gjorgieva; Kanjevac, Tatjana; Rimondini, Lia; Bosnakovski, Darko

    2016-01-01

    Understanding and apprehension of the characteristics and circumstances in which mesenchymal stem cells (MSCs) affect and make alterations (enhance or reduce) to the growth of tumors and metastasis spread is pivotal, not only for reaching the possibility to employ MSCs as drug delivery systems, but also for making forward movement in the existing knowledge of involvement of major factors (tumor microenvironment, soluble signaling molecules, etc.) in the process of carcinogenesis. This capability is reliable because MSCs present a great basis for engineering and constructions of new systems to target cancers, intended to secrete therapeutic proteins in the tumor region, or for delivering of oncolytic viruses' directly at the tumor site (targeted chemotherapy with enzyme prodrug conversion or induction of tumor cell apoptosis). MSCs as a crucial segment of the tumor surroundings and their confirmed tumor tropism, are assumed to be an open gateway for the design of promising drug delivery systems. The presented paper reviews current publications in this fieldwork, searches out the most recent patents that were published after 2012 (WO2014066122, US20140017787, WO2015100268, US20150086515), and tries to present the current progress and future prospective on the design and development in anti-cancer drug delivery systems based on MSCs.

  2. Adipose mesenchymal stem cells in the field of bone tissue engineering.

    Science.gov (United States)

    Romagnoli, Cecilia; Brandi, Maria Luisa

    2014-04-26

    Bone tissue engineering represents one of the most challenging emergent fields for scientists and clinicians. Current failures of autografts and allografts in many pathological conditions have prompted researchers to find new biomaterials able to promote bone repair or regeneration with specific characteristics of biocompatibility, biodegradability and osteoinductivity. Recent advancements for tissue regeneration in bone defects have occurred by following the diamond concept and combining the use of growth factors and mesenchymal stem cells (MSCs). In particular, a more abundant and easily accessible source of MSCs was recently discovered in adipose tissue. These adipose stem cells (ASCs) can be obtained in large quantities with little donor site morbidity or patient discomfort, in contrast to the invasive and painful isolation of bone marrow MSCs. The osteogenic potential of ASCs on scaffolds has been examined in cell cultures and animal models, with only a few cases reporting the use of ASCs for successful reconstruction or accelerated healing of defects of the skull and jaw in patients. Although these reports extend our limited knowledge concerning the use of ASCs for osseous tissue repair and regeneration, the lack of standardization in applied techniques makes the comparison between studies difficult. Additional clinical trials are needed to assess ASC therapy and address potential ethical and safety concerns, which must be resolved to permit application in regenerative medicine.

  3. A comparison of bioreactors for culture of fetal mesenchymal stem cells for bone tissue engineering.

    Science.gov (United States)

    Zhang, Zhi-Yong; Teoh, Swee Hin; Teo, Erin Yiling; Khoon Chong, Mark Seow; Shin, Chong Woon; Tien, Foo Toon; Choolani, Mahesh A; Chan, Jerry K Y

    2010-11-01

    Bioreactors provide a dynamic culture system for efficient exchange of nutrients and mechanical stimulus necessary for the generation of effective tissue engineered bone grafts (TEBG). We have shown that biaxial rotating (BXR) bioreactor-matured human fetal mesenchymal stem cell (hfMSC) mediated-TEBG can heal a rat critical sized femoral defect. However, it is not known whether optimal bioreactors exist for bone TE (BTE) applications. We systematically compared this BXR bioreactor with three most commonly used systems: Spinner Flask (SF), Perfusion and Rotating Wall Vessel (RWV) bioreactors, for their application in BTE. The BXR bioreactor achieved higher levels of cellularity and confluence (1.4-2.5x, p bioreactors operating in optimal settings. BXR bioreactor-treated scaffolds experienced earlier and more robust osteogenic differentiation on von Kossa staining, ALP induction (1.2-1.6×, p bioreactor-treated grafts, but not with the other three. BXR bioreactor enabled superior cellular proliferation, spatial distribution and osteogenic induction of hfMSC over other commonly used bioreactors. In addition, we developed and validated a non-invasive quantitative micro CT-based technique for analyzing neo-tissue formation and its spatial distribution within scaffolds.

  4. Engineered Microtissues Formed by Schiff Base Crosslinking Restore the Chondrogenic Potential of Aged Mesenchymal Stem Cells.

    Science.gov (United States)

    Millan, Christopher; Cavalli, Emma; Groth, Thomas; Maniura-Weber, Katharina; Zenobi-Wong, Marcy

    2015-06-24

    A universal method for reproducibly directing stem cell differentiation remains a major challenge for clinical applications involving cell-based therapies. The standard approach for chondrogenic induction by micromass pellet culture is highly susceptible to interdonor variability. A novel method for the fabrication of condensation-like engineered microtissues (EMTs) that utilizes hydrophilic polysaccharides to induce cell aggregation is reported here. Chondrogenesis of mesenchymal stem cells (MSCs) in EMTs is significantly enhanced compared to micromass pellets made by centrifugation measured by type II collagen gene expression, dimethylmethylene blue assay, and histology. MSCs from aged donors that fail to differentiate in pellet culture are successfully induced to synthesize cartilage-specific matrix in EMTs under identical media conditions. Furthermore, the EMT polysaccharides support the loading and release of the chondroinduction factor transforming growth factor β3 (TGF-β3). TGF-β-loaded EMTs (EMT(+TGF) ) facilitate cartilaginous tissue formation during culture in media not supplemented with the growth factor. The clinical potential of this approach is demonstrated in an explant defect model where EMT(+TGF) from aged MSCs synthesize de novo tissue containing sulfated glycosaminoglycans and type II collagen in situ.

  5. Efficient engineering of vascularized ectopic bone from human embryonic stem cell-derived mesenchymal stem cells.

    Science.gov (United States)

    Domev, Hagit; Amit, Michal; Laevsky, Ilana; Dar, Ayelet; Itskovitz-Eldor, Joseph

    2012-11-01

    Human mesenchymal stem cells (hMSCs) can be derived from various adult and fetal tissues. However, the quality of tissues for the isolation of adult and fetal hMSCs is donor dependent with a nonreproducible yield. In addition, tissue engineering and cell therapy require large-scale production of a pure population of lineage-restricted stem cells that can be easily induced to differentiate into a specific cell type. Therefore, human embryonic stem cells (hESCs) can provide an alternative, plentiful source for generation of reproducible hMSCs. We have developed efficient differentiation protocols for derivation of hMSCs from hESCs, including coculture with murine OP9 stromal cells and feeder layer-free system. Our protocols have resulted in the generation of up to 49% of hMSCs, which expressed CD105, CD90, CD29, and CD44. The hMSCs exhibited high adipogenic, chondrocytic, and osteogenic differentiation in vitro. The latter correlated with osteocalcin secretion and vascular endothelial growth factor (VEGF) production by the differentiating hMSCs. hMSC-derived osteoblasts further differentiated and formed ectopic bone in vivo, and induced the formation of blood vessels in Matrigel implants. Our protocol enables generation of a purified population of hESC-derived MSCs, with the potential of differentiating into several mesodermal lineages, and particularly into vasculogenesis-inducing osteoblasts, which can contribute to the development of bone repair protocols.

  6. Adult mesenchymal stem cells for bone and cartilage engineering: effect of scaffold materials

    Directory of Open Access Journals (Sweden)

    A Gigante

    2009-08-01

    Full Text Available Bone marrow is a useful cell source for skeletal tissue engineering approaches. In vitro differentiation of marrow mesenchymal stem cells (MSCs to chondrocytes or osteoblasts can be induced by the addition of specific growth factors to the medium. The present study evaluated the behaviour of human MSCs cultured on various scaffolds to determine whether their differentiation can be induced by cell-matrix interactions. MSCs from bone marrow collected from the acetabulum during hip arthroplasty procedures were isolated by cell sorting, expanded and characterised by a flow cytometry system. Cells were grown on three different scaffolds (type I collagen, type I + II collagen and type I collagen + hydroxyapatite membranes and analysed by histochemistry, immunohistochemistry and spectrophotometry (cell proliferation, alkaline phosphatase activity at 15 and 30 days. Widely variable cell adhesion and proliferation was observed on the three scaffolds. MSCs grown on type I+II collagen differentiated to cells expressing chondrocyte markers, while those grown on type I collagen + hydroxyapatite differentiated into osteoblast-like cells. The study highlighted that human MSCs grown on different scaffold matrices may display different behaviours in terms of cell proliferation and phenotype expression without growth factor supplementation.

  7. Full-thickness tissue engineered skin constructed with autogenic bone marrow mesenchymal stem cells

    Institute of Scientific and Technical Information of China (English)

    HE LiJuan; PEI XueTao; NAN Xue; WANG YunFang; GUAN LiDong; BAI CiXian; SHI ShuangShuang; YUAN HongFeng; CHEN Lin; LIU DaQing

    2007-01-01

    To explore the feasibility of repairing clinical cutaneous deficiency, autogenic bone marrow mesenchymal stem cells (BMSCs) were isolated and differentiated into epidermal cells and fibroblasts in vitro supplemented with different inducing factors and biomaterials to construct functional tissueengineered skin. The results showed that after 72 h induction, BMSCs displayed morphologic changes such as typical epidermal cell arrangement, from spindle shape to round or oval; tonofibrils, melanosomes and keratohyaline granules were observed under a transmission electronic microscope. The differentiated cells expressed epidermal stem cell surface marker CK19 (59.66%±4.2%) and epidermal cells differentiation marker CK10. In addition, the induced epidermal cells acquired the anti-radiation capacity featured by lowered apoptosis following exposure to UVB. On the other hand, the collagen microfibrils deposition was noticed under a transmission electronic microscope after differentiating into dermis fibroblasts; RT-PCR identified collagen type Ⅰ mRNA expression in differentiated cells;radioimmunoassay detected the secretion of interleukin-6 (IL-6) and interleukin-8 (IL-8) (up to 115.06pg/mL and 0.84 ng/mL, respectively). Further in vivo implanting BMSCs with scaffold material shortened skin wound repair significantly. In one word, autogenic BMSCs have the potential to differentiate into epidermal cells and fibroblasts in vitro, and show clinical feasibility acting as epidermis-like and dermis-like seed cells in skin engineering.

  8. Scleraxis-overexpressed human embryonic stem cell-derived mesenchymal stem cells for tendon tissue engineering with knitted silk-collagen scaffold.

    Science.gov (United States)

    Chen, Xiao; Yin, Zi; Chen, Jia-Lin; Liu, Huan-Huan; Shen, Wei-Liang; Fang, Zhi; Zhu, Ting; Ji, Junfeng; Ouyang, Hong-Wei; Zou, Xiao-Hui

    2014-06-01

    Despite our previous study that demonstrates that human embryonic stem cells (hESCs) can be used as seed cells for tendon tissue engineering after stepwise induction, suboptimal tendon regeneration implies that a new strategy needs to be developed for tendon repair. We investigated whether overexpression of the tendon-specific transcription factor scleraxis (SCX) in hESC-derived mesenchymal stem cells (hESC-MSCs) together with knitted silk-collagen sponge scaffold could promote tendon regeneration. hESCs were initially differentiated into MSCs and then engineered with scleraxis (SCX+hESC-MSCs). Engineered tendons were constructed with SCX+hESC-MSCs and a knitted silk-collagen sponge scaffold and then mechanical stress was applied. SCX elevated tendon gene expression in hESC-MSCs and concomitantly attenuated their adipogenic and chondrogenic potential. Mechanical stress further augmented the expression of tendon-specific genes in SCX+hESC-MSC-engineered tendon. Moreover, in vivo mechanical stimulation promoted the alignment of cells and increased the diameter of collagen fibers after ectopic transplantation. In the in vivo tendon repair model, the SCX+hESC-MSC-engineered tendon enhanced the regeneration process as shown by histological scores and superior mechanical performance compared with control cells, especially at early stages. Our study offers new evidence concerning the roles of SCX in tendon differentiation and regeneration. We demonstrated a novel strategy of combining hESCs, genetic engineering, and tissue-engineering principles for tendon regeneration, which are important for the future application of hESCs and silk scaffolds for tendon repair.

  9. Influence of mesenchymal stem cells on stomach tissue engineering using small intestinal submucosa.

    Science.gov (United States)

    Nakatsu, Hiroki; Ueno, Tomio; Oga, Atsunori; Nakao, Mitsuhiro; Nishimura, Taku; Kobayashi, Sei; Oka, Masaaki

    2015-03-01

    Small intestinal submucosa (SIS) is a biodegradable collagen-rich matrix containing functional growth factors. We have previously reported encouraging outcomes for regeneration of an artificial defect in the rodent stomach using SIS grafts, although the muscular layer was diminutive. In this study, we investigated the feasibility of SIS in conjunction with mesenchymal stem cells (MSCs) for regeneration of the gastrointestinal tract. MSCs from the bone marrow of green fluorescence protein (GFP)-transgenic Sprague-Dawley (SD) rats were isolated and expanded ex vivo. A 1 cm whole-layer stomach defect in SD rats was repaired using: a plain SIS graft without MSCs (group 1, control); a plain SIS graft followed by intravenous injection of MSCs (group 2); a SIS graft co-cultured with MSCs (group 3); or a SIS sandwich containing an MSC sheet (group 4). Pharmacological, electrophysiological and immunohistochemical examination was performed to evaluate the regenerated stomach tissue. Contractility in response to a muscarinic receptor agonist, a nitric oxide precursor or electrical field stimulation was observed in all groups. SIS grafts seeded with MSCs (groups 3 and 4) appeared to support improved regeneration compared with SIS grafts not seeded with MSCs (groups 1 and 2), by enabling the development of well-structured smooth muscle layers of significantly increased length. GFP expression was detected in the regenerated interstitial tissue, with fibroblast-like cells in the seeded-SIS groups. SIS potently induced pharmacological and electrophysiological regeneration of the digestive tract, and seeded MSCs provided an enriched environment that supported tissue regeneration by the SIS graft in the engineered stomach. © 2013 The Authors. Journal of Tissue Engineering and Regenerative Medicine published by John Wiley & Sons, Ltd.

  10. Age-related molecular genetic changes of murine bone marrow mesenchymal stem cells

    Directory of Open Access Journals (Sweden)

    Webster Keith A

    2010-04-01

    Full Text Available Abstract Background Mesenchymal stem cells (MSC are pluripotent cells, present in the bone marrow and other tissues that can differentiate into cells of all germ layers and may be involved in tissue maintenance and repair in adult organisms. Because of their plasticity and accessibility these cells are also prime candidates for regenerative medicine. The contribution of stem cell aging to organismal aging is under debate and one theory is that reparative processes deteriorate as a consequence of stem cell aging and/or decrease in number. Age has been linked with changes in osteogenic and adipogenic potential of MSCs. Results Here we report on changes in global gene expression of cultured MSCs isolated from the bone marrow of mice at ages 2, 8, and 26-months. Microarray analyses revealed significant changes in the expression of more than 8000 genes with stage-specific changes of multiple differentiation, cell cycle and growth factor genes. Key markers of adipogenesis including lipoprotein lipase, FABP4, and Itm2a displayed age-dependent declines. Expression of the master cell cycle regulators p53 and p21 and growth factors HGF and VEGF also declined significantly at 26 months. These changes were evident despite multiple cell divisions in vitro after bone marrow isolation. Conclusions The results suggest that MSCs are subject to molecular genetic changes during aging that are conserved during passage in culture. These changes may affect the physiological functions and the potential of autologous MSCs for stem cell therapy.

  11. Virus resistant plums through genetic engineering - from lab to market

    Science.gov (United States)

    Genetic engineering (GE) has the potential to revolutionize the genetic improvement of fruit trees and other specialty crops, to provide greater flexibility and speed in responding to changes in climate, production systems and market demands, and to maintain the competitiveness of American agricultu...

  12. Genetically modified mesenchymal stem/stromal cells transfected with adiponectin gene can stably secrete adiponectin.

    Science.gov (United States)

    Hossain, Md Murad; Murali, Malliga Raman; Kamarul, Tunku

    2017-08-01

    Mesenchymal stem/stromal cells (MSCs) hold promises for the treatment of diverse diseases and regeneration of injured tissues. Genetic modification of MSCs through gene delivery might enhance their therapeutic potential. Adiponectin has been appeared as a potential biomarker for predicting various diseases. Plasma adiponectin levels are negatively correlated with various metabolic and vascular diseases and supplementation of exogenous adiponectin ameliorates the diseases. This study aims to develop adiponectin secreting genetically modified MSCs (GM-MSCs) as a potent strategic tool to complement endogenous adiponectin for the treatment of adiponectin deficiency diseases. Human bone marrow derived MSCs were isolated, expanded in vitro and transfected with adiponectin gene containing plasmid vector. Total RNA was extracted and cDNA was prepared by reverse transcription polymerase chain reaction (RT-PCR). The expression of adiponectin gene and protein in GM-MSCs was analyzed by PCR and Western blotting respectively. The secretion of adiponectin protein from GM-MSCs was analyzed by enzyme-linked immunosorbent assay. The expression of adiponectin gene and plasmid DNA was detected in GM-MSCs but not in control group of MSCs. Adiponectin gene expression was detected in GM-MSCs at 2, 7, 14, 21 and 28days after transfection. Western blotting analysis revealed the expression of adiponectin protein only in GM-MSCs. The GM-MSCs stably secreted adiponectin protein into culture media at least for 4weeks. GM-MSCs express and secret adiponectin protein. Therefore, these adiponectin secreting GM-MSCs could be instrumental for the supplementation of adiponectin in the treatment of adiponectin deficiency related diseases. Copyright © 2017 Elsevier Inc. All rights reserved.

  13. Manipulating DNA repair for improved genetic engineering in Aspergillus

    DEFF Research Database (Denmark)

    Nødvig, Christina Spuur

    engineering strategies. Chapter 1 gives an introduction to the genus Aspergillus and some of the tools relevant to fungal genetic engineering. It also contains a short introduction to DNA repair and its interplay with gene targeting and finally an overview over the different genome editing technologies......Aspergillus is a genus of filamentous fungi, which members includes industrial producers of enzymes, organic acids and secondary metabolites, important pathogens and a model organism. As such no matter the specific area of interest there are many reasons to perform genetic engineering, whether...... it is metabolic engineering to create better performing cell factory, elucidating pathways to study secondary metabolism etc. In this thesis, the main focus is on different ways to manipulate DNA repair for optimizing gene targeting, ultimately improving the methods available for faster and better genetic...

  14. Genetic Engineering and Manufacturing of Hematopoietic Stem Cells

    Directory of Open Access Journals (Sweden)

    Xiuyan Wang

    2017-06-01

    Full Text Available The marketing approval of genetically engineered hematopoietic stem cells (HSCs as the first-line therapy for the treatment of severe combined immunodeficiency due to adenosine deaminase deficiency (ADA-SCID is a tribute to the substantial progress that has been made regarding HSC engineering in the past decade. Reproducible manufacturing of high-quality, clinical-grade, genetically engineered HSCs is the foundation for broadening the application of this technology. Herein, the current state-of-the-art manufacturing platforms to genetically engineer HSCs as well as the challenges pertaining to production standardization and product characterization are addressed in the context of primary immunodeficiency diseases (PIDs and other monogenic disorders.

  15. Enhanced genetic tools for engineering multigene traits into green algae.

    Directory of Open Access Journals (Sweden)

    Beth A Rasala

    Full Text Available Transgenic microalgae have the potential to impact many diverse biotechnological industries including energy, human and animal nutrition, pharmaceuticals, health and beauty, and specialty chemicals. However, major obstacles to sophisticated genetic and metabolic engineering in algae have been the lack of well-characterized transformation vectors to direct engineered gene products to specific subcellular locations, and the inability to robustly express multiple nuclear-encoded transgenes within a single cell. Here we validate a set of genetic tools that enable protein targeting to distinct subcellular locations, and present two complementary methods for multigene engineering in the eukaryotic green microalga Chlamydomonas reinhardtii. The tools described here will enable advanced metabolic and genetic engineering to promote microalgae biotechnology and product commercialization.

  16. Teacher-to-Teacher: An Annotated Bibliography on DNA and Genetic Engineering.

    Science.gov (United States)

    Mertens, Thomas R., Comp.

    1984-01-01

    Presented is an annotated bibliography of 24 books on DNA and genetic engineering. Areas considered in these books include: basic biological concepts to help understand advances in genetic engineering; applications of genetic engineering; social, legal, and moral issues of genetic engineering; and historical aspects leading to advances in…

  17. Epithelial-Mesenchymal Interactions in Urinary Bladder and Small Intestine and How to Apply Them in Tissue Engineering.

    Science.gov (United States)

    Jerman, Urška Dragin; Kreft, Mateja Erdani; Veranič, Peter

    2015-12-01

    Reciprocal interactions between the epithelium and mesenchyme are essential for the establishment of proper tissue morphology during organogenesis and tissue regeneration as well as for the maintenance of cell differentiation. With this review, we highlight the importance of epithelial-mesenchymal cross talk in healthy tissue and further discuss its significance in engineering functional tissues in vitro. We focus on the urinary bladder and small intestine, organs that are often compromised by disease and are as such in need of research that would advance effective treatment or tissue replacement. To date, the understanding of epithelial-mesenchymal reciprocal interactions has enabled the development of in vitro biomimetic tissue equivalents that have provided many possibilities in treating defective, damaged, or even cancerous tissues. Although research of the past several years has advanced the field of bladder and small intestine tissue engineering, one must be aware of its current limitations in successfully and above all safely introducing tissue-engineered constructs into clinical practice. Special attention is in particular needed when treating cancerous tissues, as initially successful tumor excision and tissue reconstruction may later on result in cancer recurrence due to oncogenic signals originating from an altered stroma. Recent rather poor outcomes in pioneering clinical trials of bladder reconstructions should serve as a reminder that recreating a functional organ to replace a dysfunctional one is an objective far more difficult to reach than initially foreseen. When considering effective tissue engineering approaches for diseased tissues in humans, it is imperative to introduce animal models with dysfunctional or, even more importantly, cancerous organs, which would greatly contribute to predicting possible complications and, hence, reducing risks when translating to the clinic.

  18. Exogenous enzymes upgrade transgenesis and genetic engineering of farm animals.

    Science.gov (United States)

    Bosch, Pablo; Forcato, Diego O; Alustiza, Fabrisio E; Alessio, Ana P; Fili, Alejandro E; Olmos Nicotra, María F; Liaudat, Ana C; Rodríguez, Nancy; Talluri, Thirumala R; Kues, Wilfried A

    2015-05-01

    Transgenic farm animals are attractive alternative mammalian models to rodents for the study of developmental, genetic, reproductive and disease-related biological questions, as well for the production of recombinant proteins, or the assessment of xenotransplants for human patients. Until recently, the ability to generate transgenic farm animals relied on methods of passive transgenesis. In recent years, significant improvements have been made to introduce and apply active techniques of transgenesis and genetic engineering in these species. These new approaches dramatically enhance the ease and speed with which livestock species can be genetically modified, and allow to performing precise genetic modifications. This paper provides a synopsis of enzyme-mediated genetic engineering in livestock species covering the early attempts employing naturally occurring DNA-modifying proteins to recent approaches working with tailored enzymatic systems.

  19. Response of mesenchymal stem cells to shear stress in tissue-engineered vascular grafts

    Institute of Scientific and Technical Information of China (English)

    Jian-de DONG; Yong-quan GU; Chun-min LI; Chun-ren WANG; Zeng-guo FENG; Rong-xin QIU; Bing CHEN; Jian-xin LI; Shu-wen ZHANG; Zhong-gao WANG; Jian ZHANG

    2009-01-01

    Aim: Recent studies have demonstrated that mesenchymal stem cells (MSCs) can differentiate into endothelial cells. The effect of shear stress on MSC differentiation is incompletely understood, and most studies have been based on two-dimen-sional systems. We used a model of tissue-engineered vascular grafts (TEVGs) to investigate the effects of shear stress on MSC differentiation.Methods: MSCs were isolated from canine bone marrow. The TEVG was constructed by seeding MSCs onto poly-ε-caprolactone and lactic acid (PCLA) scaffolds and subjecting them to shear stress provided by a pulsatile bioreactor for four days (two days at 1 dyne/cm2 to 15 dyne/cm2 and two days at 15 dyne/cm2).Results: Shear stress significantly increased the expression of endothelial cell markers, such as platelet-endothelial cell adhesion molecule-1 (PECAM-1), VE-cadherin, and CD34, at both the mRNA and protein levels as compared with static control cells. Protein levels of alpha-smooth muscle actin (α-SMA) and calponin were substantially reduced in shear stress-cultured cells. There was no significant change in the expression of α-SMA, smooth muscle myosin heavy chain (SMMHC)or calponin at the mRNA level.Conclusion: Shear stress upregulated the expression of endothelial cell-related markers and downregulated smooth muscle-related markers in canine MSCs. This study may serve as a basis for further investigation of the effects of shear stress on MSC differentiation in TEVGs.

  20. Calcium Phosphate Scaffolds Combined with Bone Morphogenetic Proteins or Mesenchymal Stem Cells in Bone Tissue Engineering

    Directory of Open Access Journals (Sweden)

    Han Sun

    2015-01-01

    Full Text Available Objective: The purpose of this study was to review the current status of calcium phosphate (CaP scaffolds combined with bone morphogenetic proteins (BMPs or mesenchymal stem cells (MSCs in the field of bone tissue engineering (BTE. Date Sources: Data cited in this review were obtained primarily from PubMed and Medline in publications from 1979 to 2014, with highly regarded older publications also included. The terms BTE, CaP, BMPs, and MSC were used for the literature search. Study Selection: Reviews focused on relevant aspects and original articles reporting in vitro and/or in vivo results concerning the efficiency of CaP/BMPs or CaP/MSCs composites were retrieved, reviewed, analyzed, and summarized. Results: An ideal BTE product contains three elements: Scaffold, growth factors, and stem cells. CaP-based scaffolds are popular because of their outstanding biocompatibility, bioactivity, and osteoconductivity. However, they lack stiffness and osteoinductivity. To solve this problem, composite scaffolds of CaP with BMPs have been developed. New bone formation by CaP/BMP composites can reach levels similar to those of autografts. CaP scaffolds are compatible with MSCs and CaP/MSC composites exhibit excellent osteogenesis and stiffness. In addition, a CaP/MSC/BMP scaffold can repair bone defects more effectively than an autograft. Conclusions: Novel BTE products possess remarkable osteoconduction and osteoinduction capacities, and exhibit balanced degradation with osteogenesis. Further work should yield safe, viable, and efficient materials for the repair of bone lesions.

  1. Calcium Phosphate Scaffolds Combined with Bone Morphogenetic Proteins or Mesenchymal Stem Cells in Bone Tissue Engineering

    Institute of Scientific and Technical Information of China (English)

    Han Sun; Hui-Lin Yang

    2015-01-01

    Objective:The purpose of this study was to review the current status of calcium phosphate (CaP) scaffolds combined with bone morphogenetic proteins (BMPs) or mesenchymal stem cells (MSCs) in the field of bone tissue engineering (BTE).Date Sources:Data cited in this review were obtained primarily from PubMed and Medline in publications from 1979 to 2014,with highly regarded older publications also included.The terms BTE,CaP,BMPs,and MSC were used for the literature search.Study Selection:Reviews focused on relevant aspects and original articles reporting in vitro and/or in vivo results concerning the efficiency of CaP/BMPs or CaP/MSCs composites were retrieved,reviewed,analyzed,and summarized.Results:An ideal BTE product contains three elements:Scaffold,growth factors,and stem cells.CaP-based scaffolds are popular because of their outstanding biocompatibility,bioactivity,and osteoconductivity.However,they lack stiffness and osteoinductivity.To solve this problem,composite scaffolds of CaP with BMPs have been developed.New bone formation by CaP/BMP composites can reach levels similar to those of autografts.CaP scaffolds are compatible with MSCs and CaP/MSC composites exhibit excellent osteogenesis and stiffness.In addition,a CaP/MSC/BMP scaffold can repair bone defects more effectively than an autograft.Conclusions:Novel BTE products possess remarkable osteoconduction and osteoinduction capacities,and exhibit balanced degradation with osteogenesis.Further work should yield safe,viable,and efficient materials for the repair of bone lesions.

  2. Engineering meniscus structure and function via multi-layered mesenchymal stem cell-seeded nanofibrous scaffolds.

    Science.gov (United States)

    Fisher, Matthew B; Henning, Elizabeth A; Söegaard, Nicole; Bostrom, Marc; Esterhai, John L; Mauck, Robert L

    2015-06-01

    Despite advances in tissue engineering for the knee meniscus, it remains a challenge to match the complex macroscopic and microscopic structural features of native tissue, including the circumferentially and radially aligned collagen bundles essential for mechanical function. To mimic this structural hierarchy, this study developed multi-lamellar mesenchymal stem cell (MSC)-seeded nanofibrous constructs. Bovine MSCs were seeded onto nanofibrous scaffolds comprised of poly(ε-caprolactone) with fibers aligned in a single direction (0° or 90° to the scaffold long axis) or circumferentially aligned (C). Multi-layer groups (0°/0°/0°, 90°/90°/90°, 0°/90°/0°, 90°/0°/90°, and C/C/C) were created and cultured for a total of 6 weeks under conditions favoring fibrocartilaginous tissue formation. Tensile testing showed that 0° and C single layer constructs had stiffness values several fold higher than 90° constructs. For multi-layer groups, the stiffness of 0°/0°/0° constructs was higher than all other groups, while 90°/90°/90° constructs had the lowest values. Data for collagen content showed a general positive interactive effect for multi-layers relative to single layer constructs, while a positive interaction for stiffness was found only for the C/C/C group. Collagen content and cell infiltration occurred independent of scaffold alignment, and newly formed collagenous matrix followed the scaffold fiber direction. Structural hierarchies within multi-lamellar constructs dictated biomechanical properties, and only the C/C/C constructs with non-orthogonal alignment within layers featured positive mechanical reinforcement as a consequence of the layered construction. These multi-layer constructs may serve as functional substitutes for the meniscus as well as test beds to understand the complex mechanical principles that enable meniscus function.

  3. Field Performance of a Genetically Engineered Strain of Pink Bollworm

    Science.gov (United States)

    Simmons, Gregory S.; McKemey, Andrew R.; Morrison, Neil I.; O'Connell, Sinead; Tabashnik, Bruce E.; Claus, John; Fu, Guoliang; Tang, Guolei; Sledge, Mickey; Walker, Adam S.; Phillips, Caroline E.; Miller, Ernie D.; Rose, Robert I.; Staten, Robert T.; Donnelly, Christl A.; Alphey, Luke

    2011-01-01

    Pest insects harm crops, livestock and human health, either directly or by acting as vectors of disease. The Sterile Insect Technique (SIT) – mass-release of sterile insects to mate with, and thereby control, their wild counterparts – has been used successfully for decades to control several pest species, including pink bollworm, a lepidopteran pest of cotton. Although it has been suggested that genetic engineering of pest insects provides potential improvements, there is uncertainty regarding its impact on their field performance. Discrimination between released and wild moths caught in monitoring traps is essential for estimating wild population levels. To address concerns about the reliability of current marking methods, we developed a genetically engineered strain of pink bollworm with a heritable fluorescent marker, to improve discrimination of sterile from wild moths. Here, we report the results of field trials showing that this engineered strain performed well under field conditions. Our data show that attributes critical to SIT in the field – ability to find a mate and to initiate copulation, as well as dispersal and persistence in the release area – were comparable between the genetically engineered strain and a standard strain. To our knowledge, these represent the first open-field experiments with a genetically engineered insect. The results described here provide encouragement for the genetic control of insect pests. PMID:21931649

  4. Field performance of a genetically engineered strain of pink bollworm.

    Directory of Open Access Journals (Sweden)

    Gregory S Simmons

    Full Text Available Pest insects harm crops, livestock and human health, either directly or by acting as vectors of disease. The Sterile Insect Technique (SIT--mass-release of sterile insects to mate with, and thereby control, their wild counterparts--has been used successfully for decades to control several pest species, including pink bollworm, a lepidopteran pest of cotton. Although it has been suggested that genetic engineering of pest insects provides potential improvements, there is uncertainty regarding its impact on their field performance. Discrimination between released and wild moths caught in monitoring traps is essential for estimating wild population levels. To address concerns about the reliability of current marking methods, we developed a genetically engineered strain of pink bollworm with a heritable fluorescent marker, to improve discrimination of sterile from wild moths. Here, we report the results of field trials showing that this engineered strain performed well under field conditions. Our data show that attributes critical to SIT in the field--ability to find a mate and to initiate copulation, as well as dispersal and persistence in the release area--were comparable between the genetically engineered strain and a standard strain. To our knowledge, these represent the first open-field experiments with a genetically engineered insect. The results described here provide encouragement for the genetic control of insect pests.

  5. Genetically Engineered Materials for Biofuels Production

    Science.gov (United States)

    Raab, Michael

    2012-02-01

    Agrivida, Inc., is an agricultural biotechnology company developing industrial crop feedstocks for the fuel and chemical industries. Agrivida's crops have improved processing traits that enable efficient, low cost conversion of the crops' cellulosic components into fermentable sugars. Currently, pretreatment and enzymatic conversion of the major cell wall components, cellulose and hemicellulose, into fermentable sugars is the most expensive processing step that prevents widespread adoption of biomass in biofuels processes. To lower production costs we are consolidating pretreatment and enzyme production within the crop. In this strategy, transgenic plants express engineered cell wall degrading enzymes in an inactive form, which can be reactivated after harvest. We have engineered protein elements that disrupt enzyme activity during normal plant growth. Upon exposure to specific processing conditions, the engineered enzymes are converted into their active forms. This mechanism significantly lowers pretreatment costs and enzyme loadings (>75% reduction) below those currently available to the industry.

  6. Chapter VIII. Contributions of propagation techniques and genetic modification to breeding - genetic engineering for disease resistance

    Science.gov (United States)

    Genetic engineering offers an opportunity to develop flower bulb crops with resistance to fungal, viral, and bacterial pathogens. Several of the flower bulb crops, Lilium spp., Gladiolus, Zantedeschia, Muscari, Hyacinthus, Narcissus, Ornithogalum, Iris, and Alstroemeria, have been transformed with t...

  7. Engineered Plant Minichromosome and Its Application in Genomics and Genetic Engineering

    Institute of Scientific and Technical Information of China (English)

    YU Wei-chang

    2008-01-01

    @@ Engineered minichromosomes have been constructed as novel artificial chromosome platforms for future genetic engineering in maize.We demonstrated that minichromosomes could be created by telomere-mediated chromosomal truncation of both normal A chromosomes and the supernumerary B chromosomes of maize,the minichromosomes were stable during both mitosis and meiosis,transgenes were expressed from minichromosomes,and we also demonstrated the proof of concept that minichromosomes could accept new genetic elements by a site-specific recombination system.

  8. Insights on bovine genetic engineering and cloning

    Directory of Open Access Journals (Sweden)

    Fabiana F. Bressan

    2013-12-01

    Full Text Available Transgenic technology has become an essential tool for the development of animal biotechnologies, and animal cloning through somatic cell nuclear transfer (SCNT enabled the generation of genetically modified animals utilizing previously modified and selected cell lineages as nuclei donors, assuring therefore the generation of homogeneous herds expressing the desired modification. The present study aimed to discuss the use of SCNT as an important methodology for the production of transgenic herds, and also some recent insights on genetic modification of nuclei donors and possible effects of gene induction of pluripotency on SCNT.

  9. Molecular Tissue Engineering:Applications for Modulation of Mesenchymal Stem Cells Proliferation by Transforming Growth Factor

    Institute of Scientific and Technical Information of China (English)

    GUO; Xiaodong(

    2001-01-01

    [1]郭晓东 杜靖远 郑启新等.组织工程学技术修复关节软骨缺损研究进展.国外医学生物医学工程分册 2000 23(6):330[2]郭晓东 杜靖远 郑启新等.骨关节炎基因治疗进展.国外医学老年医学分册 2001 22(1):29[3]Grande D Breitbart A Mason J et al.Cartilage tissue engineering:current limitations and solutions.Clin Orthop 1999 367S:S176[4]Brittberg M Lindahl A Nilsson A et al.Treatment of deep cartilage defects in the knee with autologous chondrocyte transplantation.N Engl J Med 1994 331:889[5]Reddi A.Morphogenesis and tissue engineering of bone and cartilage:inductive signals stem cells and biomimetic biomaterials.Tissue Eng 2000 6(4) :351[6]Freed L Martin I Vunjak-Novakovic G.Frontiers in tissue engineering:in vitro modulation of chondrogene sis.Clin Orthop 1999 367S:S46[7]Evans C Ghivizzani S Smith P et al.Using gene therapy to protect and restore cartilage.Clin Orthop 2000 379 (Suppl) :S214[8]Fernandes J Martel-Pelletier J Pelletier J.Gene therapy for osteoarthritis:new perspectives for the twentyfirst century.Clin Orthop 2000 379(Suppl):S262[9]Mason J M Breitbart A S Barcia M et al.Cartilage and bone regeneration using gene-enhanced tissue engineering.Clin Orthop 2000 379 (Suppl):S171[10]郭晓东 杜靖远 郑启新等.分子生物学在组织工程学研究中的应用前景.中华实验外科杂志 2001 18(3):283[11]郭晓东 全大萍.组织工程与生物材料.见:生物医用材料导论.李世普主编.武汉:武汉工业大学出版社 2000.302[12]Liu Y Zheng Q X Du J Y et al.Cloning and expression of rat transforming growth factorβ1 cDNA in osteoblasts.J Tongji Med Univ 2000 20:63[13]Johnstone B Yoo J.Autologous mesenchymal progenitol cells in articular cartilage repair.Clin Orthop 1999 367S:S156[14]Yoo J Mandell I Angele P et al.Chondrogenitor cells and gene therapy.Clin Orthop 2000 379 (Suppl):S164[15]Caplan A I.Mesenchymal stem cells and gene

  10. The Genetic Effects of Aging on Human Mesenchymal Stem Cells through Consecutive Subcultures

    Directory of Open Access Journals (Sweden)

    H. Pourjafari

    2013-04-01

    Full Text Available Introduction & Objective: Stem cells are determined by their unique features. One of them is high proliferation ability and the other is potency of differentiation to various tissues. The results of new works in various countries around the world including Iran show the growing momentum of works on stem cell therapy. Working with stem cells from different origins including the cells originated from fetal and adult origin (such as bone marrow, dental root, cord , etcis in progress. The goal of this work was to clear the maintenance and stability of genetic materials of human mesenchymal stem cells through consecutive subcultures in our lab conditions in Hamadan Uni-versity of Medical Sciences. Materials & Methods: In an experimental work, human umbilical cord was used as a rich source of stem cells. Isolated stem cells, after proliferation phase, increased and filled the bottom of the flasks in consecutive passages. After some successive consecutive subcultures, stem cells aged and this may cause damages to genetic material that is evaluated in primary passages (passages number 1-5, middle passages (passages number 8-12 and late passages (passages number 15-18 by the single cell gel electrophoresis assay (comet assay. Also their karyotypes were ex-amined; Solid technique and G-banding staining for chromosome analysis in the stem cells were employed. Several microscopic slides from each passage were prepared. Then 20 cells were ran-domly selected from the slides related to each group. In all selected cells, damages were exam-ined and their degree of damages were scored based on the standard patterns, from 0 to 4. The averages were compared by Kruskal–Wallis test in different groups. Results: Average scores related to three studied cell groups, primary, middle and late passages, were 0.4, 2.8 and 3.6, respectively. Prepared karyotypes from the cells belonging to passages 1-5 were normal but in the aged passages they were abnormal numerically and

  11. TMTI Task 1.6 Genetic Engineering Methods and Detection

    Energy Technology Data Exchange (ETDEWEB)

    Slezak, T; Lenhoff, R; Allen, J; Borucki, M; Vitalis, E; Gardner, S

    2009-12-04

    A large number of GE techniques can be adapted from other microorganisms to biothreat bacteria and viruses. Detection of GE in a microorganism increases in difficulty as the size of the genetic change decreases. In addition to the size of the engineered change, the consensus genomic sequence of the microorganism can impact the difficulty of detecting an engineered change in genomes that are highly variable from strain to strain. This problem will require comprehensive databases of whole genome sequences for more genetically variable biothreat bacteria and viruses. Preliminary work with microarrays for detecting synthetic elements or virulence genes and analytic bioinformatic approaches for whole genome sequence comparison to detect genetic engineering show promise for attacking this difficult problem but a large amount of future work remains.

  12. [Research progress of genetic engineering on medicinal plants].

    Science.gov (United States)

    Teng, Zhong-qiu; Shen, Ye

    2015-02-01

    The application of genetic engineering technology in modern agriculture shows its outstanding role in dealing with food shortage. Traditional medicinal plant cultivation and collection have also faced with challenges, such as lack of resources, deterioration of environment, germplasm of recession and a series of problems. Genetic engineering can be used to improve the disease resistance, insect resistance, herbicides resistant ability of medicinal plant, also can improve the medicinal plant yield and increase the content of active substances in medicinal plants. Thus, the potent biotechnology can play an important role in protection and large area planting of medicinal plants. In the development of medicinal plant genetic engineering, the safety of transgenic medicinal plants should also be paid attention to. A set of scientific safety evaluation and judgment standard which is suitable for transgenic medicinal plants should be established based on the recognition of the particularity of medicinal plants.

  13. Genetically engineered mouse models of prostate cancer

    NARCIS (Netherlands)

    Nawijn, Martijn C.; Bergman, Andreas M.; van der Poel, Henk G.

    2008-01-01

    Objectives: Mouse models of prostate cancer are used to test the contribution of individual genes to the transformation process, evaluate the collaboration between multiple genetic lesions observed in a single tumour, and perform preclinical intervention studies in prostate cancer research. Methods:

  14. GENETIC ENGINEERING OF ENHANCED MICROBIAL NITRIFICATION

    Science.gov (United States)

    Experiments were conducted to introduce genetic information in the form of antibiotic or mercuric ion resistance genes into Nitrobacter hamburgensis strain X14. The resistance genes were either stable components of broad host range plasmids or transposable genes on methods for p...

  15. Genetic engineering of sulfur-degrading Sulfolobus

    Energy Technology Data Exchange (ETDEWEB)

    Ho, N.W.Y.

    1991-01-01

    The objectives of the proposed research is to first establish a plasmid-mediated genetic transformation system for the sulfur degrading Sulfolobus, and then to clone and overexpress the genes encoding the organic-sulfur-degrading enzymes from Sulfolobus- as well as from other microorganisms, to develop a Sulfolobus-based microbial process for the removal of both organic and inorganic sulfur from coal.

  16. Mesenchymal Stromal Cells Derived from Human Umbilical Cord Tissues: Primitive Cells with Potential for Clinical and Tissue Engineering Applications

    Science.gov (United States)

    Moretti, Pierre; Hatlapatka, Tim; Marten, Dana; Lavrentieva, Antonina; Majore, Ingrida; Hass, Ralf; Kasper, Cornelia

    Mesenchymal stem or stromal cells (MSCs) have a high potential for cell-based therapies as well as for tissue engineering applications. Since Friedenstein first isolated stem or precursor cells from the human bone marrow (BM) stroma that were capable of osteogenesis, BM is currently the most common source for MSCs. However, BM presents several disadvantages, namely low frequency of MSCs, high donor-dependent variations in quality, and painful invasive intervention. Thus, tremendous research efforts have been observed during recent years to find alternative sources for MSCs.

  17. The Genetics and Biophysics of the Epithelial-Mesenchymal Transition (EMT): Can Theoretical Physics Help Cancer Biology

    Science.gov (United States)

    Levine, Herbert

    In order to spread from the primary tumor to distant sites, cancer cells must undergo a coordinated change in their phenotypic properties referred to as the ''epithelial-to-mesenchymal'' transition. We have studied the nonlinear genetic circuits that are responsible for this cellular decision-making progress and propose that the transition actually goes through a series of intermediate states. At the same time, we have formulated motility models which allow for the correlation of the state of this network and the cell's biophysical capabilities. Hopefully, these thereby efforts will help us better understand the transition to metastatic disease and possible treatments thereof.

  18. Genetic engineering of human pluripotent cells using TALE nucleases.

    Science.gov (United States)

    Hockemeyer, Dirk; Wang, Haoyi; Kiani, Samira; Lai, Christine S; Gao, Qing; Cassady, John P; Cost, Gregory J; Zhang, Lei; Santiago, Yolanda; Miller, Jeffrey C; Zeitler, Bryan; Cherone, Jennifer M; Meng, Xiangdong; Hinkley, Sarah J; Rebar, Edward J; Gregory, Philip D; Urnov, Fyodor D; Jaenisch, Rudolf

    2011-07-07

    Targeted genetic engineering of human pluripotent cells is a prerequisite for exploiting their full potential. Such genetic manipulations can be achieved using site-specific nucleases. Here we engineered transcription activator-like effector nucleases (TALENs) for five distinct genomic loci. At all loci tested we obtained human embryonic stem cell (ESC) and induced pluripotent stem cell (iPSC) clones carrying transgenic cassettes solely at the TALEN-specified location. Our data suggest that TALENs employing the specific architectures described here mediate site-specific genome modification in human pluripotent cells with similar efficiency and precision as do zinc-finger nucleases (ZFNs).

  19. Genetic program based data mining to reverse engineer digital logic

    Science.gov (United States)

    Smith, James F., III; Nguyen, Thanh Vu H.

    2006-04-01

    A data mining based procedure for automated reverse engineering and defect discovery has been developed. The data mining algorithm for reverse engineering uses a genetic program (GP) as a data mining function. A genetic program is an algorithm based on the theory of evolution that automatically evolves populations of computer programs or mathematical expressions, eventually selecting one that is optimal in the sense it maximizes a measure of effectiveness, referred to as a fitness function. The system to be reverse engineered is typically a sensor. Design documents for the sensor are not available and conditions prevent the sensor from being taken apart. The sensor is used to create a database of input signals and output measurements. Rules about the likely design properties of the sensor are collected from experts. The rules are used to create a fitness function for the genetic program. Genetic program based data mining is then conducted. This procedure incorporates not only the experts' rules into the fitness function, but also the information in the database. The information extracted through this process is the internal design specifications of the sensor. Uncertainty related to the input-output database and the expert based rule set can significantly alter the reverse engineering results. Significant experimental and theoretical results related to GP based data mining for reverse engineering will be provided. Methods of quantifying uncertainty and its effects will be presented. Finally methods for reducing the uncertainty will be examined.

  20. Enhanced Genetic Tools for Engineering Multigene Traits into Green Algae

    OpenAIRE

    Rasala, Beth A; Syh-Shiuan Chao; Matthew Pier; Daniel J Barrera; Mayfield, Stephen P.

    2014-01-01

    Transgenic microalgae have the potential to impact many diverse biotechnological industries including energy, human and animal nutrition, pharmaceuticals, health and beauty, and specialty chemicals. However, major obstacles to sophisticated genetic and metabolic engineering in algae have been the lack of well-characterized transformation vectors to direct engineered gene products to specific subcellular locations, and the inability to robustly express multiple nuclear-encoded transgenes withi...

  1. Biosynthesis and Genetic Engineering of Polyketides

    Institute of Scientific and Technical Information of China (English)

    ZHU Xiang-Cheng; WANG Qiao-Mei; SHEN Yue-Mao; DU Liang-Cheng; HUFFMAN Justin; GERBER Ryan; LOU Li-Li; XIE Yun-Xuan; LIN Ting; JORGENSON Joel; MARESCH Andrew; VOGELER Chad

    2008-01-01

    Polyketides are one of the largest groups of natural products produced by bacteria, fungi, and plants. Many of these metabolites have highly complex chemical structures and very important biological activities, including antibiotic, anticancer, immunosuppressant, and anti-cholesterol activities. In the past two decades, extensive investigations have been carried out to understand the molecular mechanisms for polyketide biosynthesis. These efforts have led to the development of various rational approaches toward engineered biosynthesis of new polyketides. More recently, the research efforts have shifted to the elucidation of the three-dimentional structure of the complex enzyme machineries for polyketide biosynthesis and to the exploitation of new sources for polyketide production, such as filamentous fungi and marine microorganisms. This review summarizes our general understanding of the biosynthetic mechanisms and the progress in engineered biosynthesis of polyketides.

  2. Nanofibers coated on acellular tissue-engineered bovine pericardium supports differentiation of mesenchymal stem cells into endothelial cells for tissue engineering.

    Science.gov (United States)

    Mathapati, Santosh; Bishi, Dillip Kumar; Venugopal, Jayarama Reddy; Cherian, Kotturathu Mammen; Guhathakurta, Soma; Ramakrishna, Seeram; Verma, Rama Shanker

    2014-04-01

    This study aimed to develop biodegradable, polymer-based nanofibers coated on acellular tissue-engineered bovine pericardium (ATEBP) for cell interfaces, enabling more exquisite functionality, such as mesenchymal stem cell (MSC) adhesion, proliferation and differentiation into endothelial cells for tissue engineering. ATEBP coated with nanofibers of poly(L-lactic acid)-co-poly(ε-caprolactone) (PLACL) and a blend of PLACL and gelatin were analyzed for human bone marrow-derived MSC adhesion, proliferation and differentiation into endothelial cells. The cell culture-based approach showed an increase in human bone marrow-derived MSC adhesion, proliferation and differentiation into endothelial cells on ATEBP coated with PLACL/gelatin nanofibers compared with ATEBP and PLACL nanofibers coated on ATEBP. ATEBP coated with PLACL/gelatin nanofibrous scaffolds, along with human bone marrow-derived MSCs differentiated into endothelial cells, might improve the scaffolds' functionality for tissue engineering.

  3. Cancer cell-oriented migration of mesenchymal stem cells engineered with an anticancer gene (PTEN: an imaging demonstration

    Directory of Open Access Journals (Sweden)

    Yang ZS

    2014-03-01

    Full Text Available Zhuo-Shun Yang,1,* Xiang-Jun Tang,2,* Xing-Rong Guo,1 Dan-Dan Zou,1 Xu-Yong Sun,3 Jing-Bo Feng,1 Jie Luo,1 Long-Jun Dai,1,4 Garth L Warnock4 1Hubei Key Laboratory of Stem Cell Research, Taihe Hospital, Hubei University of Medicine, Shiyan, People’s Republic of China; 2Department of Neurosurgery, Taihe Hospital, Hubei University of Medicine, Shiyan, People’s Republic of China; 3Guangxi Key Laboratory for Transplant Medicine, 303 Hospital of PLA, Nanning, People’s Republic of China; 4Department of Surgery, University of British Columbia, Vancouver, BC, Canada *These authors contributed equally to this work Background: Mesenchymal stem cells (MSCs have been considered to hold great potential as ideal carriers for the delivery of anticancer agents since the discovery of their tumor tropism. This study was performed to demonstrate the effects of phosphatase and tensin homolog (PTEN engineering on MSCs’ capacity for cancer cell-oriented migration. Methods: MSCs were engineered with a PTEN-bearing plasmid and the expression was confirmed with Western blotting. A human glioma cell line (DBTRG was used as the target cell; DBTRG cell-oriented migration of MSCs was monitored with a micro speed photographic system. Results: The expression of transfected PTEN in MSCs was identified by immunoblotting analysis and confirmed with cell viability assessment of target cells. The DBTRG cell-oriented migration of PTEN-engineered MSCs was demonstrated by a real-time dynamic monitoring system, and a phagocytosis-like action of MSCs was also observed. Conclusion: MSCs maintained their capacity for cancer cell-directed migration after they were engineered with anticancer genes. This study provides the first direct evidence of MSCs’ tropism post-anticancer gene engineering. Keywords: gene therapy, mesenchymal stem cells, phosphatase and tensin homolog, cancer

  4. Use of a genetically engineered mouse model as a preclinical tool for HER2 breast cancer

    Directory of Open Access Journals (Sweden)

    Helen Creedon

    2016-02-01

    Full Text Available Resistance to human epidermal growth factor receptor 2 (HER2-targeted therapies presents a major clinical problem. Although preclinical studies have identified a number of possible mechanisms, clinical validation has been difficult. This is most likely to reflect the reliance on cell-line models that do not recapitulate the complexity and heterogeneity seen in human tumours. Here, we show the utility of a genetically engineered mouse model of HER2-driven breast cancer (MMTV-NIC to define mechanisms of resistance to the pan-HER family inhibitor AZD8931. Genetic manipulation of MMTV-NIC mice demonstrated that loss of phosphatase and tensin homologue (PTEN conferred de novo resistance to AZD8931, and a tumour fragment transplantation model was established to assess mechanisms of acquired resistance. Using this approach, 50% of tumours developed resistance to AZD8931. Analysis of the resistant tumours showed two distinct patterns of resistance: tumours in which reduced membranous HER2 expression was associated with an epithelial-to-mesenchymal transition (EMT and resistant tumours that retained HER2 expression and an epithelial morphology. The plasticity of the EMT phenotype was demonstrated upon re-implantation of resistant tumours that then showed a mixed epithelial and mesenchymal phenotype. Further AZD8931 treatment resulted in the generation of secondary resistant tumours that again had either undergone EMT or retained their original epithelial morphology. The data provide a strong rationale for basing therapeutic decisions on the biology of the individual resistant tumour, which can be very different from that of the primary tumour and will be specific to individual patients.

  5. 基因工程食品%Genetic engineering food

    Institute of Scientific and Technical Information of China (English)

    汪秋安

    2003-01-01

    @@ 1 概述 近年来,生物技术在食品行业的应用迅速发展,食品生物技术包括基因工程(genetic engineering)、蛋白质工程(protein enginering)、酶工程(enzyme engineering)、发酵技术(fermentation technology)、组织与细胞培养(tissue and cell culture)、反义RNA(antisense RNA)技术等.

  6. Genetic code expansion for multiprotein complex engineering.

    Science.gov (United States)

    Koehler, Christine; Sauter, Paul F; Wawryszyn, Mirella; Girona, Gemma Estrada; Gupta, Kapil; Landry, Jonathan J M; Fritz, Markus Hsi-Yang; Radic, Ksenija; Hoffmann, Jan-Erik; Chen, Zhuo A; Zou, Juan; Tan, Piau Siong; Galik, Bence; Junttila, Sini; Stolt-Bergner, Peggy; Pruneri, Giancarlo; Gyenesei, Attila; Schultz, Carsten; Biskup, Moritz Bosse; Besir, Hueseyin; Benes, Vladimir; Rappsilber, Juri; Jechlinger, Martin; Korbel, Jan O; Berger, Imre; Braese, Stefan; Lemke, Edward A

    2016-12-01

    We present a baculovirus-based protein engineering method that enables site-specific introduction of unique functionalities in a eukaryotic protein complex recombinantly produced in insect cells. We demonstrate the versatility of this efficient and robust protein production platform, 'MultiBacTAG', (i) for the fluorescent labeling of target proteins and biologics using click chemistries, (ii) for glycoengineering of antibodies, and (iii) for structure-function studies of novel eukaryotic complexes using single-molecule Förster resonance energy transfer as well as site-specific crosslinking strategies.

  7. Successes and failures in modular genetic engineering.

    Science.gov (United States)

    Kittleson, Joshua T; Wu, Gabriel C; Anderson, J Christopher

    2012-08-01

    Synthetic biology relies on engineering concepts such as abstraction, standardization, and decoupling to develop systems that address environmental, clinical, and industrial needs. Recent advances in applying modular design to system development have enabled creation of increasingly complex systems. However, several challenges to module and system development remain, including syntactic errors, semantic errors, parameter mismatches, contextual sensitivity, noise and evolution, and load and stress. To combat these challenges, researchers should develop a framework for describing and reasoning about biological information, design systems with modularity in mind, and investigate how to predictively describe the diverse sources and consequences of metabolic load and stress.

  8. Role of nanotopography in the development of tissue engineered 3D organs and tissues using mesenchymal stem cells.

    Science.gov (United States)

    Salmasi, Shima; Kalaskar, Deepak M; Yoon, Wai-Weng; Blunn, Gordon W; Seifalian, Alexander M

    2015-03-26

    Recent regenerative medicine and tissue engineering strategies (using cells, scaffolds, medical devices and gene therapy) have led to fascinating progress of translation of basic research towards clinical applications. In the past decade, great deal of research has focused on developing various three dimensional (3D) organs, such as bone, skin, liver, kidney and ear, using such strategies in order to replace or regenerate damaged organs for the purpose of maintaining or restoring organs' functions that may have been lost due to aging, accident or disease. The surface properties of a material or a device are key aspects in determining the success of the implant in biomedicine, as the majority of biological reactions in human body occur on surfaces or interfaces. Furthermore, it has been established in the literature that cell adhesion and proliferation are, to a great extent, influenced by the micro- and nano-surface characteristics of biomaterials and devices. In addition, it has been shown that the functions of stem cells, mesenchymal stem cells in particular, could be regulated through physical interaction with specific nanotopographical cues. Therefore, guided stem cell proliferation, differentiation and function are of great importance in the regeneration of 3D tissues and organs using tissue engineering strategies. This review will provide an update on the impact of nanotopography on mesenchymal stem cells for the purpose of developing laboratory-based 3D organs and tissues, as well as the most recent research and case studies on this topic.

  9. The effect of E-beam engineered surface structures on attachment, proliferation and differentiation of human mesenchymal stem cells.

    Science.gov (United States)

    Biemond, J Elizabeth; Hannink, G; Verdonschot, N; Buma, P

    2011-01-01

    Electron beam melting (E-beam) is a new technology to produce 3-dimensional surface topographies for cementless orthopedic implants. The effect of two newly designed highly porous E-beam engineered surface structures (cubic and star) on attachment, proliferation and differentiation of human mesenchymal stem cells (hMSCs) was investigated and compared to a solid sandblasted control. SEM analysis showed that the E-beam structures allowed cells to attach and spread. Proliferation on the new surface structures was comparable to the solid control. Furthermore, differentiation on the 3D structures was comparable to the control specimen. When culturing 300,000 cells for 10 days, the cubic structure showed a significantly higher differentiation rate compared to the sandblasted specimen. We conclude that the results for attachment, proliferation and differentiation of mesenchymal stem cells on the newly engineered 3-dimensional E-beam surface topographies are promising. In vivo experiments are necessary to assess the bone ingrowth potential of the new surface structures.

  10. Genetically engineered mouse models and human osteosarcoma

    Directory of Open Access Journals (Sweden)

    Ng Alvin JM

    2012-10-01

    Full Text Available Abstract Osteosarcoma is the most common form of bone cancer. Pivotal insight into the genes involved in human osteosarcoma has been provided by the study of rare familial cancer predisposition syndromes. Three kindreds stand out as predisposing to the development of osteosarcoma: Li-Fraumeni syndrome, familial retinoblastoma and RecQ helicase disorders, which include Rothmund-Thomson Syndrome in particular. These disorders have highlighted the important roles of P53 and RB respectively, in the development of osteosarcoma. The association of OS with RECQL4 mutations is apparent but the relevance of this to OS is uncertain as mutations in RECQL4 are not found in sporadic OS. Application of the knowledge or mutations of P53 and RB in familial and sporadic OS has enabled the development of tractable, highly penetrant murine models of OS. These models share many of the cardinal features associated with human osteosarcoma including, importantly, a high incidence of spontaneous metastasis. The recent development of these models has been a significant advance for efforts to improve our understanding of the genetics of human OS and, more critically, to provide a high-throughput genetically modifiable platform for preclinical evaluation of new therapeutics.

  11. American chestnut: A test case for genetic engineering?

    Science.gov (United States)

    Leila. Pinchot

    2014-01-01

    The thought of genetically engineered (GE) trees might conjure images of mutant trees with unnatural and invasive tendencies, but there is much more to the story. GE trees are a new reality that, like it or not, will probably be part of the future of forestry. The basic inclination of most Forest Guild stewards is to reject GE trees as violating our principle to...

  12. Somatic structural rearrangements in genetically engineered mouse mammary tumors

    NARCIS (Netherlands)

    Varela, I.; Klijn, C.N.; Stephens, P.J.; Mudie, L.J.; Stebbings, L.; Galappaththige, D.; Van der Gulden, H.; Schut, E.; Klarenbeek, S.; Campbell, P.J.; Wessels, L.F.A.; Stratton, M.R.; Jonkers, J.; Futreal, P.A.; Adams, D.J.

    2010-01-01

    Background: Here we present the first paired-end sequencing of tumors from genetically engineered mouse models of cancer to determine how faithfully these models recapitulate the landscape of somatic rearrangements found in human tumors. These were models of Trp53-mutated breast cancer, Brca1- and B

  13. Intrinsic Value and the Genetic Engineering of Animals

    NARCIS (Netherlands)

    Vries, R.B.M. de

    2008-01-01

    The concept of intrinsic value is often invoked to articulate objections to the genetic engineering of animals, particularly those objections that are not directed at the negative effects the technique might have on the health and welfare of the modified animals. However, this concept was not develo

  14. A Simple Interactive Introduction to Teaching Genetic Engineering

    Science.gov (United States)

    Child, Paula

    2013-01-01

    In the UK, at key stage 4, students aged 14-15 studying GCSE Core Science or Unit 1 of the GCSE Biology course are required to be able to describe the process of genetic engineering to produce bacteria that can produce insulin. The simple interactive introduction described in this article allows students to consider the problem, devise a model and…

  15. Genetic Engineering--A Lesson on Bioethics for the Classroom.

    Science.gov (United States)

    Armstrong, Kerri; Weber, Kurt

    1991-01-01

    A unit designed to cover the topic of genetic engineering and its ethical considerations is presented. Students are expected to learn the material while using a debate format. A list of objectives for the unit, the debate format, and the results from an opinion questionnaire are described. (KR)

  16. University Students' Knowledge and Attitude about Genetic Engineering

    Science.gov (United States)

    Bal, Senol; Samanci, Nilay Keskin; Bozkurt, Orçun

    2007-01-01

    Genetic engineering and biotechnology made possible of gene transfer without discriminating microorganism, plant, animal or human. However, although these scientific techniques have benefits, they cause arguments because of their ethical and social impacts. The arguments about ethical ad social impacts of biotechnology made clear that not only…

  17. Increased production of nutriments by genetically engineered crops

    NARCIS (Netherlands)

    Sevenier, R.E.; Meer, van der I.M.; Bino, R.J.; Koops, A.J.

    2002-01-01

    Plants are the basis of human nutrition and have been selected and improved to assure this purpose. Nowadays, new technologies such as genetic engineering and genomics approaches allow further improvement of plants. We describe here three examples for which these techniques have been employed. We in

  18. De-Problematizing 'GMOs': Suggestions for Communicating about Genetic Engineering.

    Science.gov (United States)

    Blancke, Stefaan; Grunewald, Wim; De Jaeger, Geert

    2017-03-01

    The public debates concerning genetic engineering (GE) involve many non-scientific issues. The ensuing complexity is one reason why biotechnologists are reluctant to become involved. By sharing our personal experiences in science communication and suggesting ways to de-problematize GE, we aim to inspire our colleagues to engage with the public. Copyright © 2016 Elsevier Ltd. All rights reserved.

  19. Genetic engineering of syringyl-enriched lignin in plants

    Science.gov (United States)

    Chiang, Vincent Lee; Li, Laigeng

    2004-11-02

    The present invention relates to a novel DNA sequence, which encodes a previously unidentified lignin biosynthetic pathway enzyme, sinapyl alcohol dehydrogenase (SAD) that regulates the biosynthesis of syringyl lignin in plants. Also provided are methods for incorporating this novel SAD gene sequence or substantially similar sequences into a plant genome for genetic engineering of syringyl-enriched lignin in plants.

  20. Intrinsic Value and the Genetic Engineering of Animals

    NARCIS (Netherlands)

    Vries, R.B.M. de

    2008-01-01

    The concept of intrinsic value is often invoked to articulate objections to the genetic engineering of animals, particularly those objections that are not directed at the negative effects the technique might have on the health and welfare of the modified animals. However, this concept was not

  1. Effects of genetic engineering on the pharmacokinetics of antibodies

    Energy Technology Data Exchange (ETDEWEB)

    Colcher, D.; Goel, A.; Pavlinkova, G.; Beresford, G.; Booth, B.; Batra, S.K. [University of Nebraska Medical Center, Omaha NE (United States). Dept. of Pathology and Microbiology and Molecular Biology

    1999-06-01

    Monoclonal antibodies (MAbs) may be considered 'magic bullets' due to their ability to recognize and eradicate malignant cells. MAbs, however, have practical limitations for their rapid application in the clinics. The structure of the antibody molecules can be engineered to modify functional domains such as antigen-binding sites and/or effectors functions. Advanced in genetic engineering have provided rapid progress the development of new immunoglobulin constructs of MAbs with defined research and therapeutic application. Recombinant antibody constructs are being engineered, such as human mouse chimeric, domain-dispositioned, domain-deleted, humanized and single-chain Fv fragments. Genetically-engineered antibodies differ in size and rate of catabolism. Pharmacokinetics studies show that the intact IgG (150 kD), enzymatically derived fragments Fab' (50 kD) and single chain Fv (28 kD) have different clearance rates. These antibody forms clear 50% from the blood pool in 2.1 days, 30 minutes and 10 minutes, respectively. Genetically-engineered antibodies make a new class of immunotherapeutic tracers for cancer treatment.

  2. Mesenchymal stem cells delivered in a microsphere-based engineered skin contribute to cutaneous wound healing and sweat gland repair.

    Science.gov (United States)

    Huang, Sha; Lu, Gang; Wu, Yan; Jirigala, Enhe; Xu, Yongan; Ma, Kui; Fu, Xiaobing

    2012-04-01

    Bone-marrow-derived mesenchymal stem cells (BM-MSCs) can contribute to wound healing after skin injury. However, the role of BM-MSCs on repairing skin appendages in renewal tissues is incompletely explored. Moreover, most preclinical studies suggest that the therapeutic effects afforded by BM-MSCs transplantation are short-lived and relatively unstable. To assess whether engrafted bone-marrow-derived mesenchymal stem cells via a delivery system can participate in cutaneous wound healing and sweat-gland repair in mice. For safe and effective delivery of BM-MSCs to wounds, epidermal growth factor (EGF) microspheres were firstly developed to both support cells and maintain appropriate stimuli, then cell-seeded microspheres were incorporated with biomimetic scaffolds and thus fabricated an engineered skin construct with epithelial differentiation and proliferative potential. The applied efficacy was examined by implanting them into excisional wounds on both back and paws of hind legs in mice. After 3 weeks, BM-MSC-engineered skin (EGF loaded) treated wounds exhibited accelerated healing with increased re-epithelialization rates and less skin contraction. Furthermore, histological and immunofluorescence staining analysis revealed sweat glands-like structures became more apparent in BM-MSC-engineered skin (EGF loaded) treated wounds but the number of implanted BM-MSCs were decreased gradually in later phases of healing progression. Our study suggests that BM-MSCs delivered by this EGF microspheres-based engineered skin model may be a promising strategy to repair sweat glands and improve cutaneous wound healing after injury and success in this study might provide a potential benefit for BM-MSCs administration clinically. Copyright © 2012 Japanese Society for Investigative Dermatology. Published by Elsevier Ireland Ltd. All rights reserved.

  3. Targeted Eradication of Prostate Cancer Mediated by Engineered Mesenchymal Stem Cell

    Science.gov (United States)

    2007-04-01

    multiple mesenchymal tissues , such as bone, cartilage, adipose and connective tissues . Early studies demonstrated that mouse marrow stroma cell line...velocity (CBFV) using transcranial Doppler sonography (TCD), together with additional indices including salivary cortisol and subjective state. Two...without detrimental side effects to the patients. Even though the exact mechanism of preferential prostate cancer bone metastasis has not yet been well

  4. Exergetic optimization of turbofan engine with genetic algorithm method

    Energy Technology Data Exchange (ETDEWEB)

    Turan, Onder [Anadolu University, School of Civil Aviation (Turkey)], e-mail: onderturan@anadolu.edu.tr

    2011-07-01

    With the growth of passenger numbers, emissions from the aeronautics sector are increasing and the industry is now working on improving engine efficiency to reduce fuel consumption. The aim of this study is to present the use of genetic algorithms, an optimization method based on biological principles, to optimize the exergetic performance of turbofan engines. The optimization was carried out using exergy efficiency, overall efficiency and specific thrust of the engine as evaluation criteria and playing on pressure and bypass ratio, turbine inlet temperature and flight altitude. Results showed exergy efficiency can be maximized with higher altitudes, fan pressure ratio and turbine inlet temperature; the turbine inlet temperature is the most important parameter for increased exergy efficiency. This study demonstrated that genetic algorithms are effective in optimizing complex systems in a short time.

  5. Genetic engineering, a hope for sustainable biofuel production: review

    Directory of Open Access Journals (Sweden)

    Sudip Paudel

    2014-06-01

    Full Text Available The use of recently developed genetic engineering tools in combination with organisms that have the potential to produce precursors for the production of biodiesel, promises a sustainable and environment friendly energy source. Enhanced lipid production in wild type and/or genetically engineered organisms can offer sufficient raw material for industrial transesterification of plant-based triglycerides. Bio-diesel, produced with the help of genetically modified organisms, might be one of the best alternatives to fossil fuels and to mitigate various environmental hazards. DOI: http://dx.doi.org/10.3126/ije.v3i2.10644 International Journal of the Environment Vol.3(2 2014: 311-323

  6. Genetic engineering of sulfur-degrading Sulfolobus

    Energy Technology Data Exchange (ETDEWEB)

    Ho, N.W.Y. (Purdue Univ., Lafayette, IN (USA). Lab. of Renewable Resources Engineering)

    1991-01-01

    Recent studies have shown that some microorganisms can play a significant role in removing the sulfur compound from coal. Sulfolobus acidocaldarius and related species are such microorganisms. The objective of this project is to develop a genetic transformation system for Sulfolobus species so that they could become the ideal host to overproduce homologous and heterologous enzymes that are most effective for the removal of sulfur from coal, particularly organic sulfur. Last quarter, we have identified three chemicals that can inhibit the growth of S. Acidocaldarius. These chemicals can be part of the selection system for the development of a transformation system for S. acidocaldarius. Due to the fact that Sulfolobus shibatae B12 becomes increasingly more attractive as a host for housing genes encoding desulfurization enzymes, in this period we also studied the affect of these three chemicals to growth of S. shibatae B12. We found that S. shibatae B12 is also sensitive to these chemicals. This quarter we succeeded in the isolation and purification of the double-stranded DNA virus from S. shibatae B12. Furthermore, the individual EcoRI and BamH1 fragments of the virus have also been cloned into pUC19 plasmid. These plasmids will be used for the construction of the final E. coli-Sulfolobus shuttle vector. 5 Flurouracil (5FU) is one of the chemicals that inhibit growth of Sulfolobus. Resistance strain of S. acidocaldarius to 5FU has also been isolated. DNA from the 5FU resistance strain has also been isolated. 2 figs.

  7. Engineering Values Into Genetic Engineering: A Proposed Analytic Framework for Scientific Social Responsibility.

    Science.gov (United States)

    Sankar, Pamela L; Cho, Mildred K

    2015-01-01

    Recent experiments have been used to "edit" genomes of various plant, animal and other species, including humans, with unprecedented precision. Furthermore, editing the Cas9 endonuclease gene with a gene encoding the desired guide RNA into an organism, adjacent to an altered gene, could create a "gene drive" that could spread a trait through an entire population of organisms. These experiments represent advances along a spectrum of technological abilities that genetic engineers have been working on since the advent of recombinant DNA techniques. The scientific and bioethics communities have built substantial literatures about the ethical and policy implications of genetic engineering, especially in the age of bioterrorism. However, recent CRISPr/Cas experiments have triggered a rehashing of previous policy discussions, suggesting that the scientific community requires guidance on how to think about social responsibility. We propose a framework to enable analysis of social responsibility, using two examples of genetic engineering experiments.

  8. Metabolic Engineering: Techniques for analysis of targets for genetic manipulations

    DEFF Research Database (Denmark)

    Nielsen, Jens Bredal

    1998-01-01

    enzymes. Despite the prospect of obtaining major improvement through metabolic engineering, this approach is, however, not expected to completely replace the classical approach to strain improvement-random mutagenesis followed by screening. Identification of the optimal genetic changes for improvement......Metabolic engineering has been defined as the purposeful modification of intermediary metabolism using recombinant DNA techniques. With this definition metabolic engineering includes: (1) inserting new pathways in microorganisms with the aim of producing novel metabolites, e.g., production...... of polyketides by Streptomyces; (2) production of heterologous peptides, e.g., production of human insulin, erythropoitin, and tPA; and (3) improvement of both new and existing processes, e.g., production of antibiotics and industrial enzymes. Metabolic engineering is a multidisciplinary approach, which involves...

  9. Engineering genetic circuit interactions within and between synthetic minimal cells

    Science.gov (United States)

    Adamala, Katarzyna P.; Martin-Alarcon, Daniel A.; Guthrie-Honea, Katriona R.; Boyden, Edward S.

    2017-05-01

    Genetic circuits and reaction cascades are of great importance for synthetic biology, biochemistry and bioengineering. An open question is how to maximize the modularity of their design to enable the integration of different reaction networks and to optimize their scalability and flexibility. One option is encapsulation within liposomes, which enables chemical reactions to proceed in well-isolated environments. Here we adapt liposome encapsulation to enable the modular, controlled compartmentalization of genetic circuits and cascades. We demonstrate that it is possible to engineer genetic circuit-containing synthetic minimal cells (synells) to contain multiple-part genetic cascades, and that these cascades can be controlled by external signals as well as inter-liposomal communication without crosstalk. We also show that liposomes that contain different cascades can be fused in a controlled way so that the products of incompatible reactions can be brought together. Synells thus enable a more modular creation of synthetic biology cascades, an essential step towards their ultimate programmability.

  10. 76 FR 8707 - Syngenta Seeds, Inc.; Determination of Nonregulated Status for Corn Genetically Engineered To...

    Science.gov (United States)

    2011-02-15

    ..., ``Introduction of Organisms and Products Altered or Produced Through Genetic Engineering Which Are Plant Pests or... produced through genetic engineering that are plant pests or that there is reason to believe are...

  11. Genetic Engineering of Algae for Enhanced Biofuel Production ▿

    Science.gov (United States)

    Radakovits, Randor; Jinkerson, Robert E.; Darzins, Al; Posewitz, Matthew C.

    2010-01-01

    There are currently intensive global research efforts aimed at increasing and modifying the accumulation of lipids, alcohols, hydrocarbons, polysaccharides, and other energy storage compounds in photosynthetic organisms, yeast, and bacteria through genetic engineering. Many improvements have been realized, including increased lipid and carbohydrate production, improved H2 yields, and the diversion of central metabolic intermediates into fungible biofuels. Photosynthetic microorganisms are attracting considerable interest within these efforts due to their relatively high photosynthetic conversion efficiencies, diverse metabolic capabilities, superior growth rates, and ability to store or secrete energy-rich hydrocarbons. Relative to cyanobacteria, eukaryotic microalgae possess several unique metabolic attributes of relevance to biofuel production, including the accumulation of significant quantities of triacylglycerol; the synthesis of storage starch (amylopectin and amylose), which is similar to that found in higher plants; and the ability to efficiently couple photosynthetic electron transport to H2 production. Although the application of genetic engineering to improve energy production phenotypes in eukaryotic microalgae is in its infancy, significant advances in the development of genetic manipulation tools have recently been achieved with microalgal model systems and are being used to manipulate central carbon metabolism in these organisms. It is likely that many of these advances can be extended to industrially relevant organisms. This review is focused on potential avenues of genetic engineering that may be undertaken in order to improve microalgae as a biofuel platform for the production of biohydrogen, starch-derived alcohols, diesel fuel surrogates, and/or alkanes. PMID:20139239

  12. Genetic engineering of algae for enhanced biofuel production.

    Science.gov (United States)

    Radakovits, Randor; Jinkerson, Robert E; Darzins, Al; Posewitz, Matthew C

    2010-04-01

    There are currently intensive global research efforts aimed at increasing and modifying the accumulation of lipids, alcohols, hydrocarbons, polysaccharides, and other energy storage compounds in photosynthetic organisms, yeast, and bacteria through genetic engineering. Many improvements have been realized, including increased lipid and carbohydrate production, improved H(2) yields, and the diversion of central metabolic intermediates into fungible biofuels. Photosynthetic microorganisms are attracting considerable interest within these efforts due to their relatively high photosynthetic conversion efficiencies, diverse metabolic capabilities, superior growth rates, and ability to store or secrete energy-rich hydrocarbons. Relative to cyanobacteria, eukaryotic microalgae possess several unique metabolic attributes of relevance to biofuel production, including the accumulation of significant quantities of triacylglycerol; the synthesis of storage starch (amylopectin and amylose), which is similar to that found in higher plants; and the ability to efficiently couple photosynthetic electron transport to H(2) production. Although the application of genetic engineering to improve energy production phenotypes in eukaryotic microalgae is in its infancy, significant advances in the development of genetic manipulation tools have recently been achieved with microalgal model systems and are being used to manipulate central carbon metabolism in these organisms. It is likely that many of these advances can be extended to industrially relevant organisms. This review is focused on potential avenues of genetic engineering that may be undertaken in order to improve microalgae as a biofuel platform for the production of biohydrogen, starch-derived alcohols, diesel fuel surrogates, and/or alkanes.

  13. Chemical engineering of mesenchymal stem cells to induce a cell rolling response.

    Science.gov (United States)

    Sarkar, Debanjan; Vemula, Praveen Kumar; Teo, Grace S L; Spelke, Dawn; Karnik, Rohit; Wee, Le Y; Karp, Jeffrey M

    2008-11-19

    Covalently conjugated sialyl Lewis X (SLeX) on the mesenchymal stem cell (MSC) surface through a biotin-streptavidin bridge imparts leukocyte-like rolling characteristics without altering the cell phenotype and the multilineage differentiation potential. We demonstrate that the conjugation of SLeX on the MSC surface is stable, versatile, and induces a robust rolling response on P-selectin coated substrates. These results indicate the potential to increase the targeting efficiency of any cell type to specific tissue.

  14. Introduction to the application of genetic algorithms in engineering

    Directory of Open Access Journals (Sweden)

    I. S. Shaw

    1998-07-01

    Full Text Available Genetic algorithms constitute a new research area in the field of artificial intelligence. This work is aimed at their application in specific areas of engineering where good results have already been achieved. The purpose of this work is to provide a basic introduction for students as well as experienced engineers who wish to upgrade their knowledge. A distinctive feature of artificial intelligence is that instead of mathematical models, either direct human experience or certain functions of the human brain for the modelling of physical phenomena are used.

  15. The sequential seeding of epithelial and mesenchymal cells for tissue-engineered tooth regeneration.

    Science.gov (United States)

    Honda, Masaki J; Tsuchiya, Shuhei; Sumita, Yoshinori; Sagara, Hiroshi; Ueda, Minoru

    2007-02-01

    Progress is being made toward regenerating teeth by seeding dissociated postnatal odontogenic cells onto scaffolds and implanting them in vivo, but tooth morphology remains difficult to control. In this study, we aimed to facilitate tooth regeneration using a novel technique to sequentially seed epithelial cells and mesenchymal cells so that they formed appropriate interactions in the scaffold. Dental epithelium and mesenchyme from porcine third molar teeth were enzymatically separated and dissociated into single cells. Mesenchymal cells were seeded onto the surface of the scaffold and epithelial cells were then plated on top so that the two cell types were in direct contact. The cell-scaffold constructs were evaluated in vitro and also implanted into immunocompromised rats for in vivo analysis. Control groups included constructs where direct contact between the two cell types was prevented. In scaffolds seed using the novel technique, alkaline phosphatase activity was significantly greater than controls, the tooth morphology in vivo was developed in similar to that of natural tooth, and only one tooth structure formed in each scaffold. These results suggest that the novel cell-seeding technique could be useful for regulating the morphology of regenerated teeth.

  16. Vocabulary of genetic engineering. Terminology Bulletin No. 200

    Energy Technology Data Exchange (ETDEWEB)

    Delvin, E.; Pham, G.

    1990-01-01

    For some years, research, teaching and health care have been seriously affected by the lack of official terminology in the health sciences field. This vocabulary presents terminology in the field of genetic engineering, defined as those procedures arising from molecular biology which are used to manipulate DNA, the main carrier of genetic information. The vocabulary is arranged in strict alphabetical order of English terms, with cross-references to the recommended English term. These terms are accompanied by a French equivalent, followed by a context or a definition.

  17. Enhanced energy transport in genetically engineered excitonic networks

    Science.gov (United States)

    Park, Heechul; Heldman, Nimrod; Rebentrost, Patrick; Abbondanza, Luigi; Iagatti, Alessandro; Alessi, Andrea; Patrizi, Barbara; Salvalaggio, Mario; Bussotti, Laura; Mohseni, Masoud; Caruso, Filippo; Johnsen, Hannah C.; Fusco, Roberto; Foggi, Paolo; Scudo, Petra F.; Lloyd, Seth; Belcher, Angela M.

    2016-02-01

    One of the challenges for achieving efficient exciton transport in solar energy conversion systems is precise structural control of the light-harvesting building blocks. Here, we create a tunable material consisting of a connected chromophore network on an ordered biological virus template. Using genetic engineering, we establish a link between the inter-chromophoric distances and emerging transport properties. The combination of spectroscopy measurements and dynamic modelling enables us to elucidate quantum coherent and classical incoherent energy transport at room temperature. Through genetic modifications, we obtain a significant enhancement of exciton diffusion length of about 68% in an intermediate quantum-classical regime.

  18. Genetic engineering possibilities for CELSS: A bibliography and summary of techniques

    Science.gov (United States)

    Johnson, E. J.

    1982-01-01

    A bibliography of the most useful techniques employed in genetic engineering of higher plants, bacteria associated with plants, and plant cell cultures is provided. A resume of state-of-the-art genetic engineering of plants and bacteria is presented. The potential application of plant bacterial genetic engineering to CELSS (Controlled Ecological Life Support System) program and future research needs are discussed.

  19. Genetic engineering of plant food with reduced allergenicity.

    Science.gov (United States)

    Scheurer, Stephan; Sonnewald, Sophia

    2009-01-01

    Food allergies are a major health concern in industrialized countries. Since a specific immunotherapy for food allergies is not available in clinical routine praxis till now, reduction of allergens in foods, either by food processing or genetic engineering are strategies to minimize the risk of adverse reactions for food allergic patients. This review summarizes biotechnological approaches, especially the RNA interference (RNAi) technology, for the reduction of selected allergens in plant foods. So far, only a limited number of reports showing proof-of-concept of this methodology are available. Using RNAi an impressive reduction of allergen accumulation was obtained which was stable in the next generations of plants. Since threshold doses for most food allergens are not known, the beneficial effect has to be evaluated by oral challenge tests in the future. The article critically addresses the potential and limitations of genetic engineering, as well as of alternative strategies to generate "low allergic" foods.

  20. Targeted drug delivery using genetically engineered diatom biosilica.

    Science.gov (United States)

    Delalat, Bahman; Sheppard, Vonda C; Rasi Ghaemi, Soraya; Rao, Shasha; Prestidge, Clive A; McPhee, Gordon; Rogers, Mary-Louise; Donoghue, Jacqueline F; Pillay, Vinochani; Johns, Terrance G; Kröger, Nils; Voelcker, Nicolas H

    2015-11-10

    The ability to selectively kill cancerous cell populations while leaving healthy cells unaffected is a key goal in anticancer therapeutics. The use of nanoporous silica-based materials as drug-delivery vehicles has recently proven successful, yet production of these materials requires costly and toxic chemicals. Here we use diatom microalgae-derived nanoporous biosilica to deliver chemotherapeutic drugs to cancer cells. The diatom Thalassiosira pseudonana is genetically engineered to display an IgG-binding domain of protein G on the biosilica surface, enabling attachment of cell-targeting antibodies. Neuroblastoma and B-lymphoma cells are selectively targeted and killed by biosilica displaying specific antibodies sorbed with drug-loaded nanoparticles. Treatment with the same biosilica leads to tumour growth regression in a subcutaneous mouse xenograft model of neuroblastoma. These data indicate that genetically engineered biosilica frustules may be used as versatile 'backpacks' for the targeted delivery of poorly water-soluble anticancer drugs to tumour sites.

  1. Pertussis toxins, other antigens become likely targets for genetic engineering

    Energy Technology Data Exchange (ETDEWEB)

    Marwick, C.

    1990-11-14

    Genetically engineered pertussis vaccines have yet to be fully tested clinically. But early human, animal, and in vitro studies indicate effectiveness in reducing toxic effects due to Bordetella pertussis. The licensed pertussis vaccines consists of inactivated whole cells of the organism. Although highly effective, they have been associated with neurologic complications. While the evidence continues to mount that these complications are extremely rare, if they occur at all, it has affected the public's acceptance of pertussis immunization.

  2. A Quiescent, Regeneration-Responsive Tissue Engineered Mesenchymal Stem Cell Bone Marrow Niche Model via Magnetic Levitation.

    Science.gov (United States)

    Lewis, Emily Elizabeth Louise; Wheadon, Helen; Lewis, Natasha; Yang, Jingli; Mullin, Margaret; Hursthouse, Andrew; Stirling, David; Dalby, Matthew John; Berry, Catherine Cecilia

    2016-09-27

    The bone marrow niche represents a specialized environment that regulates mesenchymal stem cell quiescence and self-renewal, yet fosters stem cell migration and differentiation upon demand. An in vitro model that embodies these features would open up the ability to perform detailed study of stem cell behavior. In this paper we present a simple bone marrow-like niche model, which comprises of nanomagnetically levitated stem cells cultured as multicellular spheroids within a type I collagen gel. The stem cells maintained are nestin positive and remain quiescent until regenerative demand is placed upon them. In response to coculture wounding, they migrate and appropriately differentiate upon engraftment. This tissue engineered regeneration-responsive bone marrow-like niche model will allow for greater understanding of stem cell response to injury and also facilitate as a testing platform for drug candidates in a multiwell plate format.

  3. Genetically engineered rice. The source of β-carotene

    Directory of Open Access Journals (Sweden)

    Karol Terlecki

    2014-04-01

    Full Text Available β-carotene is a precursor of vitamin A. It is converted to vitamin A in the humans intestine by the β-carotene-15,15’-monooxygenase. Vitamin A is essential to support vision, as an antioxidant it protects the body from free radicals, it helps to integrate the immune system, as well as takes part in cellular differentiation and proliferation. Vitamin A deficiency is a major public health problem especially among developing countries. Nyctalopia, commonly known as „Night Blindness” is one of the major symptoms of Vitamin A deficiency (VAD. Plants such as apricots, broccoli, carrots, and sweet potatoes are rich in β-carotene. Some of the plants are characterized by a higher content of provitamin-A. Among vegetables rich sources of β-carotene are: carrots, pumpkin, spinach, lettuce, green peas, tomatoes, watercress, broccoli and parsley leaves. Amongst fruits the highest content of β-carotene is in apricot, cherry, sweet cherry, plum, orange and mango. The aim of the present study was to analyze available literature data of increasing the content of β-carotene in genetically engineered rice. The genetically modified cultivar contains additional genes: PSY and CRTI thanks to which rice seed endosperm contains β-carotene. Genetically engineered rice with β-carotene is an effective source of vitamin A, it contains approximately 30 μg β-carotene per 1 g. Fortunately some of the advantages of Genetically Modified Food give an opportunity to reduce VAD worldwide, by introducing the rice which has been genetically engineered to be rich in β-carotene. The popularity of this plant as an element of nutrition is simultaneously a source of vitamin A.

  4. Unraveling the neurobiology of nicotine dependence using genetically engineered mice.

    Science.gov (United States)

    Stoker, Astrid K; Markou, Athina

    2013-08-01

    This review article provides an overview of recent studies of nicotine dependence and withdrawal that used genetically engineered mice. Major progress has been made in recent years with mutant mice that have knockout and gain-of-function of specific neuronal nicotinic acetylcholine receptor (nAChR) subunit genes. Nicotine exerts its actions by binding to neuronal nAChRs that consist of five subunits. The different nAChR subunits that combine to compose a receptor determine the distinct pharmacological and kinetic properties of the specific nAChR. Recent findings in genetically engineered mice have indicated that while α4-containing and β2-containing nAChRs are involved in the acquisition of nicotine self-administration and initial stages of nicotine dependence, α7 homomeric nAChRs appear to be involved in the later stages of nicotine dependence. In the medial habenula, α5-containing, α3-containing, and β4-containing nAChRs were shown to be crucially important in the regulation of the aversive aspects of nicotine. Studies of the involvement of α6 nAChR subunits in nicotine dependence have only recently emerged. The use of genetically engineered mice continues to vastly improve our understanding of the neurobiology of nicotine dependence and withdrawal.

  5. Versatile RNA-sensing transcriptional regulators for engineering genetic networks.

    Science.gov (United States)

    Lucks, Julius B; Qi, Lei; Mutalik, Vivek K; Wang, Denise; Arkin, Adam P

    2011-05-24

    The widespread natural ability of RNA to sense small molecules and regulate genes has become an important tool for synthetic biology in applications as diverse as environmental sensing and metabolic engineering. Previous work in RNA synthetic biology has engineered RNA mechanisms that independently regulate multiple targets and integrate regulatory signals. However, intracellular regulatory networks built with these systems have required proteins to propagate regulatory signals. In this work, we remove this requirement and expand the RNA synthetic biology toolkit by engineering three unique features of the plasmid pT181 antisense-RNA-mediated transcription attenuation mechanism. First, because the antisense RNA mechanism relies on RNA-RNA interactions, we show how the specificity of the natural system can be engineered to create variants that independently regulate multiple targets in the same cell. Second, because the pT181 mechanism controls transcription, we show how independently acting variants can be configured in tandem to integrate regulatory signals and perform genetic logic. Finally, because both the input and output of the attenuator is RNA, we show how these variants can be configured to directly propagate RNA regulatory signals by constructing an RNA-meditated transcriptional cascade. The combination of these three features within a single RNA-based regulatory mechanism has the potential to simplify the design and construction of genetic networks by directly propagating signals as RNA molecules.

  6. Promising Therapeutic Strategies for Mesenchymal Stem Cell-Based Cardiovascular Regeneration: From Cell Priming to Tissue Engineering

    Directory of Open Access Journals (Sweden)

    Seung Taek Ji

    2017-01-01

    Full Text Available The primary cause of death among chronic diseases worldwide is ischemic cardiovascular diseases, such as stroke and myocardial infarction. Recent evidence indicates that adult stem cell therapies involving cardiovascular regeneration represent promising strategies to treat cardiovascular diseases. Owing to their immunomodulatory properties and vascular repair capabilities, mesenchymal stem cells (MSCs are strong candidate therapeutic stem cells for use in cardiovascular regeneration. However, major limitations must be overcome, including their very low survival rate in ischemic lesion. Various attempts have been made to improve the poor survival and longevity of engrafted MSCs. In order to develop novel therapeutic strategies, it is necessary to first identify stem cell modulators for intracellular signal triggering or niche activation. One promising therapeutic strategy is the priming of therapeutic MSCs with stem cell modulators before transplantation. Another is a tissue engineering-based therapeutic strategy involving a cell scaffold, a cell-protein-scaffold architecture made of biomaterials such as ECM or hydrogel, and cell patch- and 3D printing-based tissue engineering. This review focuses on the current clinical applications of MSCs for treating cardiovascular diseases and highlights several therapeutic strategies for promoting the therapeutic efficacy of MSCs in vitro or in vivo from cell priming to tissue engineering strategies, for use in cardiovascular regeneration.

  7. Influence of culture conditions and extracellular matrix alignment on human mesenchymal stem cells invasion into decellularized engineered tissues.

    Science.gov (United States)

    Weidenhamer, Nathan K; Moore, Dusty L; Lobo, Fluvio L; Klair, Nathaniel T; Tranquillo, Robert T

    2015-05-01

    The variables that influence the in vitro recellularization potential of decellularized engineered tissues, such as cell culture conditions and scaffold alignment, have yet to be explored. The goal of this work was to explore the influence of insulin and ascorbic acid and extracellular matrix (ECM) alignment on the recellularization of decellularized engineered tissue by human mesenchymal stem cells (hMSCs). Aligned and non-aligned tissues were created by specifying the geometry and associated mechanical constraints to fibroblast-mediated fibrin gel contraction and remodelling using circular and C-shaped moulds. Decellularized tissues (matrices) of the same alignment were created by decellularization with detergents. Ascorbic acid promoted the invasion of hMSCs into the matrices due to a stimulated increase in motility and proliferation. Invasion correlated with hyaluronic acid secretion, α-smooth muscle actin expression and decreased matrix thickness. Furthermore, hMSCs invasion into aligned and non-aligned matrices was not different, although there was a difference in cell orientation. Finally, we show that hMSCs on the matrix surface appear to differentiate toward a smooth muscle cell or myofibroblast phenotype with ascorbic acid treatment. These results inform the strategy of recellularizing decellularized engineered tissue with hMSCs.

  8. Bioprinting Organotypic Hydrogels with Improved Mesenchymal Stem Cell Remodeling and Mineralization Properties for Bone Tissue Engineering.

    Science.gov (United States)

    Duarte Campos, Daniela Filipa; Blaeser, Andreas; Buellesbach, Kate; Sen, Kshama Shree; Xun, Weiwei; Tillmann, Walter; Fischer, Horst

    2016-06-01

    3D-manufactured hydrogels with precise contours and biological adhesion motifs are interesting candidates in the regenerative medicine field for the culture and differentiation of human bone-marrow-derived mesenchymal stem cells (MSCs). 3D-bioprinting is a powerful technique to approach one step closer the native organization of cells. This study investigates the effect of the incorporation of collagen type I in 3D-bioprinted polysaccharide-based hydrogels to the modulation of cell morphology, osteogenic remodeling potential, and mineralization. By combining thermo-responsive agarose hydrogels with collagen type I, the mechanical stiffness and printing contours of printed constructs can be improved compared to pure collagen hydrogels which are typically used as standard materials for MSC osteogenic differentiation. The results presented here show that MSC not only survive the 3D-bioprinting process but also maintain the mesenchymal phenotype, as proved by live/dead staining and immunocytochemistry (vimentin positive, CD34 negative). Increased solids concentrations of collagen in the hydrogel blend induce changes in cell morphology, namely, by enhancing cell spreading, that ultimately contribute to enhanced and directed MSC osteogenic differentiation. 3D-bioprinted agarose-collagen hydrogels with high-collagen ratio are therefore feasible for MSC osteogenic differentiation, contrarily to low-collagen blends, as proved by two-photon microscopy, Alizarin Red staining, and real-time polymerase chain reaction.

  9. The delicate balance in genetically engineering live vaccines.

    Science.gov (United States)

    Galen, James E; Curtiss, Roy

    2014-07-31

    Contemporary vaccine development relies less on empirical methods of vaccine construction, and now employs a powerful array of precise engineering strategies to construct immunogenic live vaccines. In this review, we will survey various engineering techniques used to create attenuated vaccines, with an emphasis on recent advances and insights. We will further explore the adaptation of attenuated strains to create multivalent vaccine platforms for immunization against multiple unrelated pathogens. These carrier vaccines are engineered to deliver sufficient levels of protective antigens to appropriate lymphoid inductive sites to elicit both carrier-specific and foreign antigen-specific immunity. Although many of these technologies were originally developed for use in Salmonella vaccines, application of the essential logic of these approaches will be extended to development of other enteric vaccines where possible. A central theme driving our discussion will stress that the ultimate success of an engineered vaccine rests on achieving the proper balance between attenuation and immunogenicity. Achieving this balance will avoid over-activation of inflammatory responses, which results in unacceptable reactogenicity, but will retain sufficient metabolic fitness to enable the live vaccine to reach deep tissue inductive sites and trigger protective immunity. The breadth of examples presented herein will clearly demonstrate that genetic engineering offers the potential for rapidly propelling vaccine development forward into novel applications and therapies which will significantly expand the role of vaccines in public health.

  10. Genetic Engineering In BioButanol Production And Tolerance

    Directory of Open Access Journals (Sweden)

    Ashok Rao

    Full Text Available ABSTRACT The growing need to address current energy and environmental problems has sparked an interest in developing improved biological methods to produce liquid fuels from renewable sources. Higher-chain alcohols possess chemical properties that are more similar to gasoline. Ethanol and butanol are two products which are used as biofuel. Butanol production was more concerned than ethanol because of its high octane number. Unfortunately, these alcohols are not produced efficiently in natural microorganisms, and thus economical production in industrial volumes remains a challenge. The synthetic biology, however, offers additional tools to engineer synthetic pathways in user-friendly hosts to help increase titers and productivity of bio-butanol. Knock out and over-expression of genes is the major approaches towards genetic manipulation and metabolic engineering of microbes. Yet there are TargeTron Technology, Antisense RNA and CRISPR technology has a vital role in genome manipulation of C.acetobutylicum. This review concentrates on the recent developments for efficient production of butanol and butanol tolerance by various genetically engineered microbes.

  11. Placenta Derived Mesenchymal Stem Cells Hosted on RKKP Glass-Ceramic: A Tissue Engineering Strategy for Bone Regenerative Medicine Applications

    Directory of Open Access Journals (Sweden)

    Mario Ledda

    2016-01-01

    Full Text Available In tissue engineering protocols, the survival of transplanted stem cells is a limiting factor that could be overcome using a cell delivery matrix able to support cell proliferation and differentiation. With this aim, we studied the cell-friendly and biocompatible behavior of RKKP glass-ceramic coated Titanium (Ti surface seeded with human amniotic mesenchymal stromal cells (hAMSCs from placenta. The sol-gel synthesis procedure was used to prepare the RKKP glass-ceramic material, which was then deposited onto the Ti surface by Pulsed Laser Deposition method. The cell metabolic activity and proliferation rate, the cytoskeletal actin organization, and the cell cycle phase distribution in hAMSCs seeded on the RKKP coated Ti surface revealed no significant differences when compared to the cells grown on the treated plastic Petri dish. The health of of hAMSCs was also analysed studying the mRNA expressions of MSC key genes and the osteogenic commitment capability using qRT-PCR analysis which resulted in being unchanged in both substrates. In this study, the combination of the hAMSCs’ properties together with the bioactive characteristics of RKKP glass-ceramics was investigated and the results obtained indicate its possible use as a new and interesting cell delivery system for bone tissue engineering and regenerative medicine applications.

  12. Placenta Derived Mesenchymal Stem Cells Hosted on RKKP Glass-Ceramic: A Tissue Engineering Strategy for Bone Regenerative Medicine Applications

    Science.gov (United States)

    Fosca, Marco; De Bonis, Angela; Curcio, Mariangela; Lolli, Maria Grazia; De Stefanis, Adriana; Marchese, Rodolfo; Rau, Julietta V.

    2016-01-01

    In tissue engineering protocols, the survival of transplanted stem cells is a limiting factor that could be overcome using a cell delivery matrix able to support cell proliferation and differentiation. With this aim, we studied the cell-friendly and biocompatible behavior of RKKP glass-ceramic coated Titanium (Ti) surface seeded with human amniotic mesenchymal stromal cells (hAMSCs) from placenta. The sol-gel synthesis procedure was used to prepare the RKKP glass-ceramic material, which was then deposited onto the Ti surface by Pulsed Laser Deposition method. The cell metabolic activity and proliferation rate, the cytoskeletal actin organization, and the cell cycle phase distribution in hAMSCs seeded on the RKKP coated Ti surface revealed no significant differences when compared to the cells grown on the treated plastic Petri dish. The health of of hAMSCs was also analysed studying the mRNA expressions of MSC key genes and the osteogenic commitment capability using qRT-PCR analysis which resulted in being unchanged in both substrates. In this study, the combination of the hAMSCs' properties together with the bioactive characteristics of RKKP glass-ceramics was investigated and the results obtained indicate its possible use as a new and interesting cell delivery system for bone tissue engineering and regenerative medicine applications. PMID:28078286

  13. TRAIL-engineered pancreas-derived mesenchymal stem cells: characterization and cytotoxic effects on pancreatic cancer cells.

    Science.gov (United States)

    Moniri, M R; Sun, X-Y; Rayat, J; Dai, D; Ao, Z; He, Z; Verchere, C B; Dai, L-J; Warnock, G L

    2012-09-01

    Mesenchymal stem cells (MSCs) have attracted great interest in cancer therapy owing to their tumor-oriented homing capacity and the feasibility of autologous transplantation. Currently, pancreatic cancer patients face a very poor prognosis, primarily due to the lack of therapeutic strategies with an effective degree of specificity. Anticancer gene-engineered MSCs specifically target tumor sites and can produce anticancer agents locally and constantly. This study was performed to characterize pancreas-derived MSCs and investigate the effects of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-engineered MSCs on pancreatic cancer cells under different culture conditions. Pancreas-derived MSCs exhibited positive expression on CD44, CD73, CD95, CD105, negative on CD34 and differentiated into adipogenic and osteogenic cells. TRAIL expression was assessed by both enzyme-linked immunosorbent assay and western blot analysis. Different patterns of TRAIL receptor expression were observed on the pancreatic cancer cell lines, including PANC1, HP62, ASPC1, TRM6 and BXPC3. Cell viability was assessed using a real-time monitoring system. Pancreatic cancer cell death was proportionally related to conditioned media from MSC(nsTRAIL) and MSC(stTRAIL). The results suggest that MSCs exhibit intrinsic inhibition of pancreatic cancer cells and that this effect can be potentiated by TRAIL-transfection on death receptor-bearing cell types.

  14. Neural Differentiation of Mesenchymal Stem Cells on Scaffolds for Nerve Tissue Engineering Applications.

    Science.gov (United States)

    Quintiliano, Kerlin; Crestani, Thayane; Silveira, Davi; Helfer, Virginia Etges; Rosa, Annelise; Balbueno, Eduardo; Steffens, Daniela; Jotz, Geraldo Pereira; Pilger, Diogo André; Pranke, Patricia

    2016-11-01

    Scaffolds produced by electrospinning act as supports for cell proliferation and differentiation, improved through the release of neurotrophic factors. The objective of this study was to develop aligned and random nanofiber scaffolds with and without nerve growth factor to evaluate the potential of mesenchymal stem cells (MSCs) for neural differentiation. Nanofiber morphology, diameter, degradability, cell morphology, adhesion, proliferation, viability, cytotoxicity, and neural differentiation were performed to characterize the scaffolds. The expression for nestin, β-III tubulin, and neuron-specific enolase was also evaluated. The scaffolds demonstrated a satisfactory environment for MSC growth, being nontoxic. The MSCs cultivated on the scaffolds were able to adhere and proliferate. The evaluation of neural differentiation indicated that in all groups of scaffolds the MSCs were able to upregulate neural gene expression.

  15. Engineering structures and functions of mesenchymal stem cells by suspended large-area graphene nanopatterns

    Science.gov (United States)

    Kim, Jangho; Bae, Won-Gyu; Park, Subeom; Kim, Yeon Ju; Jo, Insu; Park, Sunho; Li Jeon, Noo; Kwak, Woori; Cho, Seoae; Park, Jooyeon; Kim, Hong Nam; Choi, Kyoung Soon; Seonwoo, Hoon; Choung, Yun-Hoon; Choung, Pill-Hoon; Hong, Byung Hee; Chung, Jong Hoon

    2016-09-01

    Inspired by the hierarchical nanofibrous and highly oriented structures of natural extracellular matrices, we report a rational design of chemical vapor deposition graphene-anchored scaffolds that provide both physical and chemical cues in a multilayered organization to control the adhesion and functions of cells for regenerative medicine. These hierarchical platforms are fabricated by transferring large graphene film onto nanogroove patterns. The top graphene layer exhibits planar morphology with slight roughness (∼20 nm between peaks) due to the underlying topography, which results in a suspended structure between the nanoridges. We demonstrate that the adhesion and differentiation of human mesenchymal stem cells were sensitively controlled and enhanced by the both the nanotopography and graphene cues in our scaffolds. Our results indicate that the layered physical and chemical cues can affect the apparent cell behaviors, and can synergistically enhance cell functionality. Therefore, these suspended graphene platforms may be used to advance regenerative medicine.

  16. Process engineering of high voltage alginate encapsulation of mesenchymal stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Gryshkov, Oleksandr, E-mail: gryshkov@imp.uni-hannover.de [Institute for Multiphase Processes, Leibniz University Hannover, D-30167 Hannover (Germany); Pogozhykh, Denys, E-mail: pogozhykh@imp.uni-hannover.de [Institute for Multiphase Processes, Leibniz University Hannover, D-30167 Hannover (Germany); Zernetsch, Holger, E-mail: zernetsch@imp.uni-hannover.de [Institute for Multiphase Processes, Leibniz University Hannover, D-30167 Hannover (Germany); Hofmann, Nicola, E-mail: hofmann@imp.uni-hannover.de [Institute for Multiphase Processes, Leibniz University Hannover, D-30167 Hannover (Germany); Mueller, Thomas, E-mail: mueller.thomas@mh-hannover.de [Institute for Transfusion Medicine, Medical School Hannover, D-30625 Hannover (Germany); Glasmacher, Birgit, E-mail: glasmacher@imp.uni-hannover.de [Institute for Multiphase Processes, Leibniz University Hannover, D-30167 Hannover (Germany)

    2014-03-01

    Encapsulation of stem cells in alginate beads is promising as a sophisticated drug delivery system in treatment of a wide range of acute and chronic diseases. However, common use of air flow encapsulation of cells in alginate beads fails to produce beads with narrow size distribution, intact spherical structure and controllable sizes that can be scaled up. Here we show that high voltage encapsulation (≥ 15 kV) can be used to reproducibly generate spherical alginate beads (200–400 μm) with narrow size distribution (± 5–7%) in a controlled manner under optimized process parameters. Flow rate of alginate solution ranged from 0.5 to 10 ml/h allowed producing alginate beads with a size of 320 and 350 μm respectively, suggesting that this approach can be scaled up. Moreover, we found that applied voltages (15–25 kV) did not alter the viability and proliferation of encapsulated mesenchymal stem cells post-encapsulation and cryopreservation as compared to air flow. We are the first who employed a comparative analysis of electro-spraying and air flow encapsulation to study the effect of high voltage on alginate encapsulated cells. This report provides background in application of high voltage to encapsulate living cells for further medical purposes. Long-term comparison and work on alginate–cell interaction within these structures will be forthcoming. - Highlights: • High voltage alginate encapsulation of mesenchymal stem cells (MSCs) was designed. • Reproducible and spherical alginate beads were generated via high voltage. • Air flow encapsulation was utilized as a comparative approach to high voltage. • High voltage did not alter the viability and proliferation of encapsulated MSCs. • High voltage encapsulation can be scaled up and applied in cell-based therapy.

  17. Multipotent mesenchymal stem cells from human subacromial bursa: potential for cell based tendon tissue engineering.

    Science.gov (United States)

    Song, Na; Armstrong, April D; Li, Feng; Ouyang, Hongsheng; Niyibizi, Christopher

    2014-01-01

    Rotator cuff injuries are a common clinical problem either as a result of overuse or aging. Biological approaches to tendon repair that involve use of scaffolding materials or cell-based approaches are currently being investigated. The cell-based approaches are focused on applying multipotent mesenchymal stem cells (MSCs) mostly harvested from bone marrow. In the present study, we focused on characterizing cells harvested from tissues associated with rotator cuff tendons based on an assumption that these cells would be more appropriate for tendon repair. We isolated MSCs from bursa tissue associated with rotator cuff tendons and characterized them for multilineage differentiation in vitro and in vivo. Human bursa was obtained from patients undergoing rotator cuff surgery and cells within were isolated using collagenase and dispase digestion. The cells isolated from the tissues were characterized for osteoblastic, adipogenic, chondrogenic, and tenogenic differentiation in vitro and in vivo. The results showed that the cells isolated from bursa tissue exhibited MSCs characteristics as evidenced by the expression of putative cell surface markers attributed to MSCs. The cells exhibited high proliferative capacity and differentiated toward cells of mesenchymal lineages with high efficiency. Bursa-derived cells expressed markers of tenocytes when treated with bone morphogenetic protein-12 (BMP-12) and assumed aligned morphology in culture. Bursa cells pretreated with BMP-12 and seeded in ceramic scaffolds formed extensive bone, as well as tendon-like tissue in vivo. Bone formation was demonstrated by histological analysis and immunofluorescence for DMP-1 in tissue sections made from the scaffolds seeded with the cells. Tendon-like tissue formed in vivo consisted of parallel collagen fibres typical of tendon tissues. Bursa-derived cells also formed a fibrocartilagenous tissue in the ceramic scaffolds. Taken together, the results demonstrate a new source of MSCs with a

  18. Genetic engineering of fibrous proteins: spider dragline silk and collagen.

    Science.gov (United States)

    Wong Po Foo, Cheryl; Kaplan, David L

    2002-10-18

    Various strategies have been employed to genetically engineer fibrous proteins. Two examples, the subject of this review, include spider dragline silk from Nephila clavipes and collagen. These proteins are highlighted because of their unique mechanical and biological properties related to controlled release, biomaterials and tissue engineering. Cloning and expression of native genes and synthetic artificial variants of the consensus sequence repeats from the native genes has been accomplished. Expression of recombinant silk and collagen proteins has been reported in a variety of host systems, including bacteria, yeast, insect cells, plants and mammalian cells. Future utility for these proteins for biomedical materials is expected to increase as needs expand for designer materials with tailored mechanical properties and biological interactions to elicit specific responses in vitro and in vivo.

  19. Strategies to genetically engineer T cells for cancer immunotherapy.

    Science.gov (United States)

    Spear, Timothy T; Nagato, Kaoru; Nishimura, Michael I

    2016-06-01

    Immunotherapy is one of the most promising and innovative approaches to treat cancer, viral infections, and other immune-modulated diseases. Adoptive immunotherapy using gene-modified T cells is an exciting and rapidly evolving field. Exploiting knowledge of basic T cell biology and immune cell receptor function has fostered innovative approaches to modify immune cell function. Highly translatable clinical technologies have been developed to redirect T cell specificity by introducing designed receptors. The ability to engineer T cells to manifest desired phenotypes and functions is now a thrilling reality. In this review, we focus on outlining different varieties of genetically engineered T cells, their respective advantages and disadvantages as tools for immunotherapy, and their promise and drawbacks in the clinic.

  20. Cancer Regression in Patients After Transfer of Genetically Engineered Lymphocytes

    Science.gov (United States)

    Morgan, Richard A.; Dudley, Mark E.; Wunderlich, John R.; Hughes, Marybeth S.; Yang, James C.; Sherry, Richard M.; Royal, Richard E.; Topalian, Suzanne L.; Kammula, Udai S.; Restifo, Nicholas P.; Zheng, Zhili; Nahvi, Azam; de Vries, Christiaan R.; Rogers-Freezer, Linda J.; Mavroukakis, Sharon A.; Rosenberg, Steven A.

    2006-10-01

    Through the adoptive transfer of lymphocytes after host immunodepletion, it is possible to mediate objective cancer regression in human patients with metastatic melanoma. However, the generation of tumor-specific T cells in this mode of immunotherapy is often limiting. Here we report the ability to specifically confer tumor recognition by autologous lymphocytes from peripheral blood by using a retrovirus that encodes a T cell receptor. Adoptive transfer of these transduced cells in 15 patients resulted in durable engraftment at levels exceeding 10% of peripheral blood lymphocytes for at least 2 months after the infusion. We observed high sustained levels of circulating, engineered cells at 1 year after infusion in two patients who both demonstrated objective regression of metastatic melanoma lesions. This study suggests the therapeutic potential of genetically engineered cells for the biologic therapy of cancer.

  1. Genetically modified cells in regenerative medicine and tissue engineering.

    Science.gov (United States)

    Sheyn, Dima; Mizrahi, Olga; Benjamin, Shimon; Gazit, Zulma; Pelled, Gadi; Gazit, Dan

    2010-06-15

    Regenerative medicine appears to take as its patron, the Titan Prometheus, whose liver was able to regenerate daily, as the field attempts to restore lost, damaged, or aging cells and tissues. The tremendous technological progress achieved during the last decade in gene transfer methods and imaging techniques, as well as recent increases in our knowledge of cell biology, have opened new horizons in the field of regenerative medicine. Genetically engineered cells are a tool for tissue engineering and regenerative medicine, albeit a tool whose development is fraught with difficulties. Gene-and-cell therapy offers solutions to severe problems faced by modern medicine, but several impediments obstruct the path of such treatments as they move from the laboratory toward the clinical setting. In this review we provide an overview of recent advances in the gene-and-cell therapy approach and discuss the main hurdles and bottlenecks of this approach on its path to clinical trials and prospective clinical practice.

  2. Advances in Research on Genetically Engineered Plants for Metal Resistance

    Institute of Scientific and Technical Information of China (English)

    Ri-Qing Zhang; Chun-Fang Tang; Shi-Zhi Wen; Yun-Guo Liu; Ke-Lin Li

    2006-01-01

    The engineering application of natural hyperaccumulators in removing or inactivating metal pollutants from soil and surface water in field trials mostly presents the insurmountable shortcoming of low efficiency owing to their little biomass and slow growth. Based on further understanding of the molecular mechanism of metal uptake, translocation, and also the separation, identification, and cloning of some related functional genes, this article highlights and summarizes in detail the advances in research on transgenic techniques, such as Agrobacterium tumefaciens-mediated transformation and particle bombardment, in breeding of plants for metal resistance and accumulation, and points out that deepening the development of transgenic plants is one of the efficient approaches to improving phytoremediation efficiency of metal-contaminated environments. From the viewpoint of sustainable development, governments should strengthen support to the development of genetic engineering for metal resistance and accumulation in plants.

  3. Hairy Root and Its Application in Plant Genetic Engineering

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    Agrobacterium rhizogenes Conn. causes hairy root disease in plants. Hairy root-infected A. rhizogenes is characterized by a high growth rate and genetic stability. Hairy root cultures have been proven to be an efficient means of producing secondary metabolites that are normally biosynthesized in roots of differentiated plants.Furthermore, a transgenic root system offers tremendous potential for introducing additional genes along with the Ri plasmid, especially with modified genes, into medicinal plant cells with A. rhizogenes vector systems.The cultures have turned out to be a valuable tool with which to study the biochemical properties and the gene expression profile of metabolic pathways. Moreover, the cultures can be used to elucidate the intermediates and key enzymes involved in the biosynthesis of secondary metabolites. The present article discusses various applications of hairy root cultures in plant genetic engineering and potential problems associated with them.

  4. Engineering Genetically-Encoded Mineralization and Magnetism via Directed Evolution.

    Science.gov (United States)

    Liu, Xueliang; Lopez, Paola A; Giessen, Tobias W; Giles, Michael; Way, Jeffrey C; Silver, Pamela A

    2016-11-29

    Genetically encoding the synthesis of functional nanomaterials such as magnetic nanoparticles enables sensitive and non-invasive biological sensing and control. Via directed evolution of the natural iron-sequestering ferritin protein, we discovered key mutations that lead to significantly enhanced cellular magnetism, resulting in increased physical attraction of ferritin-expressing cells to magnets and increased contrast for cellular magnetic resonance imaging (MRI). The magnetic mutants further demonstrate increased iron biomineralization measured by a novel fluorescent genetic sensor for intracellular free iron. In addition, we engineered Escherichia coli cells with multiple genomic knockouts to increase cellular accumulation of various metals. Lastly to explore further protein candidates for biomagnetism, we characterized members of the DUF892 family using the iron sensor and magnetic columns, confirming their intracellular iron sequestration that results in increased cellular magnetization.

  5. Genetic-evolution-based optimization methods for engineering design

    Science.gov (United States)

    Rao, S. S.; Pan, T. S.; Dhingra, A. K.; Venkayya, V. B.; Kumar, V.

    1990-01-01

    This paper presents the applicability of a biological model, based on genetic evolution, for engineering design optimization. Algorithms embodying the ideas of reproduction, crossover, and mutation are developed and applied to solve different types of structural optimization problems. Both continuous and discrete variable optimization problems are solved. A two-bay truss for maximum fundamental frequency is considered to demonstrate the continuous variable case. The selection of locations of actuators in an actively controlled structure, for minimum energy dissipation, is considered to illustrate the discrete variable case.

  6. Biodegradation of azo dyes by genetically engineered azoreductase

    Institute of Scientific and Technical Information of China (English)

    WANG Jing; YAN Bin; ZHOU Ji-ti; BAO Yong-ming; LU Hong; YUAN Xiao-dong

    2005-01-01

    A azoreductase gene with 537 bp was obtained by PGR amplification from Rhodobacter sphaeroides AS1.1737. The enzyme,with a molecular weight of 18.7 kD, was efficiently expressed in Escherichia coli and its biodegradation characteristics for azo dyes were investigated. Furthermore, the reaction kinetics and mechanism of azo dyes catalyzed by the genetically engineered azoreductase were studied in detail. The presence of a hydrazo-intermediate was identified, which provided a convincing evidence for the assumption that azo dyes were degraded via an incomplete reduction stage.

  7. Genetic-evolution-based optimization methods for engineering design

    Science.gov (United States)

    Rao, S. S.; Pan, T. S.; Dhingra, A. K.; Venkayya, V. B.; Kumar, V.

    1990-01-01

    This paper presents the applicability of a biological model, based on genetic evolution, for engineering design optimization. Algorithms embodying the ideas of reproduction, crossover, and mutation are developed and applied to solve different types of structural optimization problems. Both continuous and discrete variable optimization problems are solved. A two-bay truss for maximum fundamental frequency is considered to demonstrate the continuous variable case. The selection of locations of actuators in an actively controlled structure, for minimum energy dissipation, is considered to illustrate the discrete variable case.

  8. Comparison of drug and cell-based delivery: engineered adult mesenchymal stem cells expressing soluble tumor necrosis factor receptor II prevent arthritis in mouse and rat animal models.

    Science.gov (United States)

    Liu, Linda N; Wang, Gang; Hendricks, Kyle; Lee, Keunmyoung; Bohnlein, Ernst; Junker, Uwe; Mosca, Joseph D

    2013-05-01

    Rheumatoid arthritis (RA) is a systemic autoimmune disease with unknown etiology where tumor necrosis factor-α (TNFα) plays a critical role. Etanercept, a recombinant fusion protein of human soluble tumor necrosis factor receptor II (hsTNFR) linked to the Fc portion of human IgG1, is used to treat RA based on the rationale that sTNFR binds TNFα and blocks TNFα-mediated inflammation. We compared hsTNFR protein delivery from genetically engineered human mesenchymal stem cells (hMSCs) with etanercept. Blocking TNFα-dependent intercellular adhesion molecule-1 expression on transduced hMSCs and inhibition of nitric oxide production from TNFα-treated bovine chondrocytes by conditioned culture media from transduced hMSCs demonstrated the functionality of the hsTNFR construction. Implanted hsTNFR-transduced mesenchymal stem cells (MSCs) reduced mouse serum circulating TNFα generated from either implanted TNFα-expressing cells or lipopolysaccharide induction more effectively than etanercept (TNFα, 100%; interleukin [IL]-1α, 90%; and IL-6, 60% within 6 hours), suggesting faster clearance of the soluble tumor necrosis factor receptor (sTNFR)-TNFα complex from the animals. In vivo efficacy of sTNFR-transduced MSCs was illustrated in two (immune-deficient and immune-competent) arthritic rodent models. In the antibody-induced arthritis BalbC/SCID mouse model, intramuscular injection of hsTNFR-transduced hMSCs reduced joint inflammation by 90% compared with untransduced hMSCs; in the collagen-induced arthritis Fischer rat model, both sTNFR-transduced rat MSCs and etanercept inhibited joint inflammation by 30%. In vitro chondrogenesis assays showed the ability of TNFα and IL1α, but not interferon γ, to inhibit hMSC differentiation to chondrocytes, illustrating an additional negative role for inflammatory cytokines in joint repair. The data support the utility of hMSCs as therapeutic gene delivery vehicles and their potential to be used in alleviating inflammation

  9. 76 FR 5780 - Determination of Regulated Status of Alfalfa Genetically Engineered for Tolerance to the...

    Science.gov (United States)

    2011-02-02

    ... Animal and Plant Health Inspection Service Determination of Regulated Status of Alfalfa Genetically... regulated status of alfalfa genetically engineered for tolerance to the herbicide glyphosate based on APHIS... decision and determination on the petition regarding the regulated status of alfalfa genetically engineered...

  10. Efficient derivation and genetic modifications of human pluripotent stem cells on engineered human feeder cell lines.

    Science.gov (United States)

    Zou, Chunlin; Chou, Bin-Kuan; Dowey, Sarah N; Tsang, Kitman; Huang, Xiaosong; Liu, Cyndi F; Smith, Cory; Yen, Jonathan; Mali, Prashant; Zhang, Yu Alex; Cheng, Linzhao; Ye, Zhaohui

    2012-08-10

    Derivation of pluripotent stem cells (iPSCs) induced from somatic cell types and the subsequent genetic modifications of disease-specific or patient-specific iPSCs are crucial steps in their applications for disease modeling as well as future cell and gene therapies. Conventional procedures of these processes require co-culture with primary mouse embryonic fibroblasts (MEFs) to support self-renewal and clonal growth of human iPSCs as well as embryonic stem cells (ESCs). However, the variability of MEF quality affects the efficiencies of all these steps. Furthermore, animal sourced feeders may hinder the clinical applications of human stem cells. In order to overcome these hurdles, we established immortalized human feeder cell lines by stably expressing human telomerase reverse transcriptase, Wnt3a, and drug resistance genes in adult mesenchymal stem cells. Here, we show that these immortalized human feeders support efficient derivation of virus-free, integration-free human iPSCs and long-term expansion of human iPSCs and ESCs. Moreover, the drug-resistance feature of these feeders also supports nonviral gene transfer and expression at a high efficiency, mediated by piggyBac DNA transposition. Importantly, these human feeders exhibit superior ability over MEFs in supporting homologous recombination-mediated gene targeting in human iPSCs, allowing us to efficiently target a transgene into the AAVS1 safe harbor locus in recently derived integration-free iPSCs. Our results have great implications in disease modeling and translational applications of human iPSCs, as these engineered human cell lines provide a more efficient tool for genetic modifications and a safer alternative for supporting self-renewal of human iPSCs and ESCs.

  11. Development of 3D in vitro platform technology to engineer mesenchymal stem cells

    OpenAIRE

    Hosseinkhani H; Hong P; Yu D; Chen Y.; Ickowicz D; Farber I; Domb AJ

    2012-01-01

    Hossein Hosseinkhani,1 Po-Da Hong,1 Dah-Shyong Yu,2 Yi-Ru Chen,3 Diana Ickowicz,4 Ira-Yudovin Farber,4 Abraham J Domb41Graduate Institute of Biomedical Engineering, National Taiwan University of Science and Technology (TAIWANTECH), 2Nanomedicine Research Center, National Defense Medical Center, Taipei, Taiwan, 3Department of Biomedical Engineering, National Yang-Ming University, Taipei, Taiwan, 4Institute of Drug Research, The Center for Nanoscience and Nanotechnology, School of Pharmacy-Facu...

  12. Rhizobia species: A Boon for "Plant Genetic Engineering".

    Science.gov (United States)

    Patel, Urmi; Sinha, Sarika

    2011-10-01

    Since past three decades new discoveries in plant genetic engineering have shown remarkable potentials for crop improvement. Agrobacterium Ti plasmid based DNA transfer is no longer the only efficient way of introducing agronomically important genes into plants. Recent studies have explored a novel plant genetic engineering tool, Rhizobia sp., as an alternative to Agrobacterium, thereby expanding the choice of bacterial species in agricultural plant biotechnology. Rhizobia sp. serve as an open license source with no major restrictions in plant biotechnology and help broaden the spectrum for plant biotechnologists with respect to the use of gene transfer vehicles in plants. New efficient transgenic plants can be produced by transferring genes of interest using binary vector carrying Rhizobia sp. Studies focusing on the interactions of Rhizobia sp. with their hosts, for stable and transient transformation and expression of genes, could help in the development of an adequate gene transfer vehicle. Along with being biologically beneficial, it may also bring a new means for fast economic development of transgenic plants, thus giving rise to a new era in plant biotechnology, viz. "Rhizobia mediated transformation technology."

  13. Study on biofortification of rice by targeted genetic engineering

    Directory of Open Access Journals (Sweden)

    Sumon M. Hossain

    2012-12-01

    Full Text Available Micronutrient malnutrition is a major health problem in Bangladesh and also in many other developing countries, where a diversified diet is not affordable for the majority. In the present world- one, out of seven people suffers from hunger. Yet, there is a stealthier form of hunger than lack of food: micronutrient malnutrition or hidden hunger. While often providing enough calories, monotonous diets (of rural poor frequently fail to deliver sufficient quantities of essential minerals and vitamins. Due to micronutrient deficiencies different characteristic features have been observed to the victims. Various estimates indicate that over two-thirds of the world population, for the most part women and children specially, pre-school children are deficient in at least one micronutrient. This can have devastating consequences for the life, health and well being of the individuals concerned (like premature death, blindness, weakened immune systems etc. Genetic engineering approach is the upcoming strategy to solve this problem. Genetically engineered biofortified staple crops specially, rice that are high in essential micronutrients (Fe, Zn, vitamin A and adapted to local growing environments have the potential to significantly reduce the prevalence of micronutrient deficiencies specially to the rural poor.

  14. Genetic engineering and chemical conjugation of potato virus X.

    Science.gov (United States)

    Lee, Karin L; Uhde-Holzem, Kerstin; Fischer, Rainer; Commandeur, Ulrich; Steinmetz, Nicole F

    2014-01-01

    Here we report the genetic engineering and chemical modification of potato virus X (PVX) for the presentation of various peptides, proteins, and fluorescent dyes, or other chemical modifiers. Three different ways of genetic engineering are described and by these means, peptides are successfully expressed not only when the foot and mouth disease virus (FMDV) 2A sequence or a flexible glycine-serine linker is included, but also when the peptide is fused directly to the PVX coat protein. When larger proteins or unfavorable peptide sequences are presented, a partial fusion via the FMDV 2A sequence is preferable. When these PVX chimeras retain the ability to assemble into viral particles and are thus able to infect plants systemically, they can be utilized to inoculate susceptible plants for isolation of sufficient amounts of virus particles for subsequent chemical modification. Chemical modification is required for the display of nonbiological ligands such as fluorophores, polymers, and small drug compounds. We present three methods of chemical bioconjugation. For direct conjugation of small chemical modifiers to solvent exposed lysines, N-hydroxysuccinimide chemistry can be applied. Bio-orthogonal reactions such as copper-catalyzed azide-alkyne cycloaddition or hydrazone ligation are alternatives to achieve more efficient conjugation (e.g., when working with high molecular weight or insoluble ligands). Furthermore, hydrazone ligation offers an attractive route for the introduction of pH-cleavable cargos (e.g., therapeutic molecules).

  15. Process engineering of high voltage alginate encapsulation of mesenchymal stem cells.

    Science.gov (United States)

    Gryshkov, Oleksandr; Pogozhykh, Denys; Zernetsch, Holger; Hofmann, Nicola; Mueller, Thomas; Glasmacher, Birgit

    2014-03-01

    Encapsulation of stem cells in alginate beads is promising as a sophisticated drug delivery system in treatment of a wide range of acute and chronic diseases. However, common use of air flow encapsulation of cells in alginate beads fails to produce beads with narrow size distribution, intact spherical structure and controllable sizes that can be scaled up. Here we show that high voltage encapsulation (≥ 15 kV) can be used to reproducibly generate spherical alginate beads (200-400 μm) with narrow size distribution (± 5-7%) in a controlled manner under optimized process parameters. Flow rate of alginate solution ranged from 0.5 to 10 ml/h allowed producing alginate beads with a size of 320 and 350 μm respectively, suggesting that this approach can be scaled up. Moreover, we found that applied voltages (15-25 kV) did not alter the viability and proliferation of encapsulated mesenchymal stem cells post-encapsulation and cryopreservation as compared to air flow. We are the first who employed a comparative analysis of electro-spraying and air flow encapsulation to study the effect of high voltage on alginate encapsulated cells. This report provides background in application of high voltage to encapsulate living cells for further medical purposes. Long-term comparison and work on alginate-cell interaction within these structures will be forthcoming.

  16. Prediction of jet engine parameters for control design using genetic programming

    OpenAIRE

    Martínez-Arellano, G; Cant, R; Nolle, L

    2014-01-01

    The simulation of a jet engine behavior is widely used in many different aspects of the engine development and maintenance. Achieving high quality jet engine control systems requires the iterative use of these simulations to virtually test the performance of the engine avoiding any possible damage on the real engine. Jet engine simulations involve the use of mathematical models which are complex and may not always be available. This paper introduces an approach based on Genetic Programming (G...

  17. Biosynthetic Studies and Genetic Engineering of Pactamycin Analogs with Improved Selectivity toward Malarial Parasites

    National Research Council Canada - National Science Library

    Lu, Wanli; Roongsawang, Niran; Mahmud, Taifo

    2011-01-01

    .... However, through extensive biosynthetic studies and genetic engineering, we were able to produce analogs of pactamycin that show potent antimalarial activity, but lack significant antibacterial...

  18. Fibronectin-Alginate microcapsules improve cell viability and protein secretion of encapsulated Factor IX-engineered human mesenchymal stromal cells.

    Science.gov (United States)

    Sayyar, Bahareh; Dodd, Megan; Marquez-Curtis, Leah; Janowska-Wieczorek, Anna; Hortelano, Gonzalo

    2015-01-01

    Continuous delivery of proteins by engineered cells encapsu-lated in biocompatible polymeric microcapsules is of considerable therapeutic potential. However, this technology has not lived up to expectations due to inadequate cell--matrix interactions and subsequent cell death. In this study we hypoth-esize that the presence of fibronectin in an alginate matrix may enhance the viability and functionality of encapsulated human cord blood-derived mesenchymal stromal cells (MSCs) expressing the human Factor IX (FIX) gene. MSCs were encapsulated in alginate-PLL microcapsules containing 10, 100, or 500 μg/ml fibronectin to ameliorate cell survival. MSCs in microcapsules with 100 and 500 μg/ml fibronectin demonstrated improved cell viability and proliferation and higher FIX secretion compared to MSCs in non-supplemented microcapsules. In contrast, 10 μg/ml fibronectin did not significantly affect the viability and protein secretion from the encapsulated cells. Differentiation studies demonstrated osteogenic (but not chondrogenic or adipogenic) differentiation capability and efficient FIX secretion of the enclosed MSCs in the fibronectin-alginate suspension culture. Thus, the use of recombinant MSCs encapsulated in fibronectin-alginate microcapsules in basal or osteogenic cultures may be of practical use in the treatment of hemophilia B.

  19. Retina tissue engineering by conjunctiva mesenchymal stem cells encapsulated in fibrin gel: Hypotheses on novel approach to retinal diseases treatment.

    Science.gov (United States)

    Soleimannejad, Mostafa; Ebrahimi-Barough, Somayeh; Nadri, Samad; Riazi-Esfahani, Mohammad; Soleimani, Masoud; Tavangar, Seyed Mohammad; Ai, Jafar

    2017-04-01

    Retinitis pigmentosa (RP) and age related macular degeneration (AMD) are two retinal diseases that progress by photoreceptor cells death. In retinal transplantation studies, stem and progenitor cells inject into the sub retinal space or vitreous and then these cells can be migrate to the site of retinal degeneration and locate in the host retina and restitute vision. Our hypothesis suggests that using human conjunctiva stem cells (as the source for increasing the number of human stem cells progenitor cells in retina dysfunction diseases) with fibrin gel and also assessing its relating in vitro (cellular and molecular processes) and in vivo (vision tests and pathology) could be a promising strategy for treatment of AMD and RP disorders. In this idea, we describe a novel approach for retina tissue engineering with differentiation of conjunctiva mesenchymal stem cells (CJMSCs) into photoreceptor-like cells in fibrin gel with induction medium contain taurine. For assessment of differentiation, immunocytochemistry and real time PCR are used for the expression of Rhodopsin, RPE65, Nestin as differentiated photoreceptor cell markers in 2D and 3D culture. The results show that fibrin gel will offer a proper 3D scaffold for CJMSCs derived photoreceptor cell-like cells. Application of immune-privileged, readily available sources of adult stem cells like human conjunctiva stem cells with fibrin gel would be a promising strategy to increase the number of photoreceptor progenitor cells and promote involuntary angiogenesis needed in retina layer repair and regeneration. Copyright © 2017 Elsevier Ltd. All rights reserved.

  20. Tissue engineering with peripheral blood-derived mesenchymal stem cells promotes the regeneration of injured peripheral nerves.

    Science.gov (United States)

    Pan, Mengjie; Wang, Xianghai; Chen, Yijing; Cao, Shangtao; Wen, Jinkun; Wu, Guofeng; Li, Yuanyuan; Li, Lixia; Qian, Changhui; Qin, Zhenqi; Li, Zhenlin; Tan, Dandan; Fan, Zhihao; Wu, Wutian; Guo, Jiasong

    2017-06-01

    Peripheral nerve injury repair can be enhanced by Schwann cell (SC) transplantation, but clinical applications are limited by the lack of a cell source. Thus, alternative systems for generating SCs are desired. Herein, we found the peripheral blood-derived mesenchymal stem cells (PBMSCs) could be induced into SC like cells with expressing SC-specific markers (S100, P75NTR and CNPase) and functional factors (NGF, NT-3, c-Fos, and Krox20). When the induced PBMSCs (iPBMSCs) were transplanted into crushed rat sciatic nerves, they functioned as SCs by wrapping the injured axons and expressing myelin specific marker of MBP. Furthermore, iPBMSCs seeded in an artificial nerve conduit to bridge a 10-mm defect in a sciatic nerve achieved significant nerve regeneration outcomes, including axonal regeneration and remyelination, nerve conduction recovery, and restoration of motor function, and attenuated myoatrophy and neuromuscular junction degeneration in the target muscle. Overall, the data from this study indicated that PBMSCs can transdifferentiate towards SC-like cells and have potential as grafting cells for nerve tissue engineering. Copyright © 2017. Published by Elsevier Inc.

  1. Cryo-chemical decellularization of the whole liver for mesenchymal stem cells-based functional hepatic tissue engineering.

    Science.gov (United States)

    Jiang, Wei-Cheng; Cheng, Yu-Hao; Yen, Meng-Hua; Chang, Yin; Yang, Vincent W; Lee, Oscar K

    2014-04-01

    Liver transplantation is the ultimate treatment for severe hepatic failure to date. However, the limited supply of donor organs has severely hampered this treatment. So far, great potentials of using mesenchymal stem cells (MSCs) to replenish the hepatic cell population have been shown; nevertheless, there still is a lack of an optimal three-dimensional scaffold for generation of well-transplantable hepatic tissues. In this study, we utilized a cryo-chemical decellularization method which combines physical and chemical approach to generate acellular liver scaffolds (ALS) from the whole liver. The produced ALS provides a biomimetic three-dimensional environment to support hepatic differentiation of MSCs, evidenced by expression of hepatic-associated genes and marker protein, glycogen storage, albumin secretion, and urea production. It is also found that hepatic differentiation of MSCs within the ALS is much more efficient than two-dimensional culture in vitro. Importantly, the hepatic-like tissues (HLT) generated by repopulating ALS with MSCs are able to act as functional grafts and rescue lethal hepatic failure after transplantation in vivo. In summary, the cryo-chemical method used in this study is suitable for decellularization of liver and create acellular scaffolds that can support hepatic differentiation of MSCs and be used to fabricate functional tissue-engineered liver constructs.

  2. Mechanical stimulation of mesenchymal stem cells: Implications for cartilage tissue engineering.

    Science.gov (United States)

    Fahy, Niamh; Alini, Mauro; Stoddart, Martin J

    2017-08-01

    Articular cartilage is a load-bearing tissue playing a crucial mechanical role in diarthrodial joints, facilitating joint articulation, and minimizing wear. The significance of biomechanical stimuli in the development of cartilage and maintenance of chondrocyte phenotype in adult tissues has been well documented. Furthermore, dysregulated loading is associated with cartilage pathology highlighting the importance of mechanical cues in cartilage homeostasis. The repair of damaged articular cartilage resulting from trauma or degenerative joint disease poses a major challenge due to a low intrinsic capacity of cartilage for self-renewal, attributable to its avascular nature. Bone marrow-derived mesenchymal stem cells (MSCs) are considered a promising cell type for cartilage replacement strategies due to their chondrogenic differentiation potential. Chondrogenesis of MSCs is influenced not only by biological factors but also by the environment itself, and various efforts to date have focused on harnessing biomechanics to enhance chondrogenic differentiation of MSCs. Furthermore, recapitulating mechanical cues associated with cartilage development and homeostasis in vivo, may facilitate the development of a cellular phenotype resembling native articular cartilage. The goal of this review is to summarize current literature examining the effect of mechanical cues on cartilage homeostasis, disease, and MSC chondrogenesis. The role of biological factors produced by MSCs in response to mechanical loading will also be examined. An in-depth understanding of the impact of mechanical stimulation on the chondrogenic differentiation of MSCs in terms of endogenous bioactive factor production and signaling pathways involved, may identify therapeutic targets and facilitate the development of more robust strategies for cartilage replacement using MSCs. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res. © 2017 Orthopaedic Research Society

  3. Clinical application of human mesenchymal stromal cells for bone tissue engineering

    NARCIS (Netherlands)

    Ganguly, Anindita; Meijer, Gert; van Blitterswijk, Clemens; de Boer, Jan

    2010-01-01

    The gold standard in the repair of bony defects is autologous bone grafting, even though it has drawbacks in terms of availability and morbidity at the harvesting site. Bone-tissue engineering, in which osteogenic cells and scaffolds are combined, is considered as a potential bone graft substitute

  4. Engineered myocardial tissues constructed in vivo using cardiomyocyte-like cells derived from bone marrow mesenchymal stem cells in rats

    Directory of Open Access Journals (Sweden)

    Xing Yujie

    2012-01-01

    Full Text Available Abstract Background To explore the feasibility of constructing engineered myocardial tissues (EMTs in vivo, using polylactic acid -co-glycolic acid (PLGA for scaffold and cardiomyocyte-like cells derived from bone marrow mesenchymal stem cells (BMMSCs for seeded cells. Methods BMMSCs were isolated from femur and tibia of Sprague-Dawley (SD rats by density-gradient centrifugation. The third passage cells were treated with 10 μmol/L 5-azacytidine (5-aza and 0.1 μmol/L angiotensin II (Ang II for 24 h, followed by culturing in complete medium for 3 weeks to differentiated into cardiomyocyte-like cells. The cardiomyocyte-like cells were seeded into PLGA scaffolds to form the grafts. The grafts were cultured in the incubator for three days and then implanted into the peritoneal cavity of SD rats. Four weeks later, routine hematoxylin-eosin (HE staining, immunohistochemical staining for myocardium-specific cardiac troponin I (cTnI, scanning electron microscopy and transmission electron microscopy were used to analyze the morphology and microconstruction of the EMTs in host rats. Results HE staining showed that the cardiomyocyte-like cells distributed equally in the PLGA scaffold, and the nuclei arranged in the spindle shape. Immunohistochemical staining revealed that majority of engrafted cells in the PLGA -Cardiomyocyte-like cells group were positive for cTnI. Scanning electron microscopy showed that the inoculated cells well attached to PLGA and grew in 3 dimensions in construct. Transmission electron microscopy showed that the EMTs contained well arranged myofilaments paralleled to the longitudinal cell axis, the cells were rich in endoplasmic reticulum and mitochondria, while desmosomes, gap junction and Z line-like substances were also can be observed as well within the engrafted cells. Conclusion We have developed an in vivo method to construct engineered myocardial tissue. The in vivo microenvironment helped engrafted cells/tissue survive and

  5. Bone tissue engineering via human induced pluripotent, umbilical cord and bone marrow mesenchymal stem cells in rat cranium.

    Science.gov (United States)

    Wang, Ping; Liu, Xian; Zhao, Liang; Weir, Michael D; Sun, Jirun; Chen, Wenchuan; Man, Yi; Xu, Hockin H K

    2015-05-01

    Human induced pluripotent stem cells (hiPSCs) are an exciting cell source with great potential for tissue engineering. Human bone marrow mesenchymal stem cells (hBMSCs) have been used in clinics but are limited by several disadvantages, hence alternative sources of MSCs such as umbilical cord MSCs (hUCMSCs) are being investigated. However, there has been no report comparing hiPSCs, hUCMSCs and hBMSCs for bone regeneration. The objectives of this pilot study were to investigate hiPSCs, hUCMSCs and hBMSCs for bone tissue engineering, and compare their bone regeneration via seeding on biofunctionalized macroporous calcium phosphate cement (CPC) in rat cranial defects. For all three types of cells, approximately 90% of the cells remained alive on CPC scaffolds. Osteogenic genes were up-regulated, and mineral synthesis by cells increased with time in vitro for all three types of cells. The new bone area fractions at 12weeks (mean±sd; n=6) were (30.4±5.8)%, (27.4±9.7)% and (22.6±4.7)% in hiPSC-MSC-CPC, hUCMSC-CPC and hBMSC-CPC respectively, compared to (11.0±6.3)% for control (pcells (p>0.1). New blood vessel density was higher in cell-seeded groups than control (pcells was confirmed via immunohistochemical staining. In conclusion, (1) hiPSCs, hUCMSCs and hBMSCs greatly enhanced bone regeneration, more than doubling the new bone amount of cell-free CPC control; (2) hiPSC-MSCs and hUCMSCs represented viable alternatives to hBMSCs; (3) biofunctionalized macroporous CPC-stem cell constructs had a robust capacity for bone regeneration. Published by Elsevier Ltd.

  6. Biology of lung cancer: genetic mutation, epithelial-mesenchymal transition, and cancer stem cells.

    Science.gov (United States)

    Aoi, Takashi

    2016-09-01

    At present, most cases of unresectable cancer cannot be cured. Genetic mutations, EMT, and cancer stem cells are three major issues linked to poor prognosis in such cases, all connected by inter- and intra-tumor heterogeneity. Issues on inter-/intra-tumor heterogeneity of genetic mutation could be resolved with recent and future technologies of deep sequencers, whereas, regarding such issues as the "same genome, different epigenome/phenotype", we expect to solve many of these problems in the future through further research in stem cell biology. We herein review and discuss the three major issues in the biology of cancers, especially from the standpoint of stem cell biology.

  7. Combination therapy with leflunomide and genetic engineering biological agents

    Directory of Open Access Journals (Sweden)

    Nataliya Vladimirovna Chichasova

    2011-06-01

    Full Text Available The paper gives data on the use of a combination of genetic engineering biological agents (GEBAs and leflunomide in patients with rheumatoid arthritis (RA. In accordance with the international guidelines, the majority of GEBAs should be given in a combination with methotrexate (MTX, which increases the efficacy of a number of GEBAs (tumor necrosis factor-α inhibitors, rituximab and affects tolerability (remikeid, humira. However, MTX cannot be always used in real practice. The data given in the paper on the efficiency and safety of the coadministration of leflunomide and a GEBA in patients with active RA, which are based on the results of randomized studies and national registers, including the Russian one, point to the compatibility of the results of treatment with this and GEBA-MTX combinations.

  8. Optochemical control of genetically engineered neuronal nicotinic acetylcholine receptors

    Science.gov (United States)

    Tochitsky, Ivan; Banghart, Matthew R.; Mourot, Alexandre; Yao, Jennifer Z.; Gaub, Benjamin; Kramer, Richard H.; Trauner, Dirk

    2012-02-01

    Advances in synthetic chemistry, structural biology, molecular modelling and molecular cloning have enabled the systematic functional manipulation of transmembrane proteins. By combining genetically manipulated proteins with light-sensitive ligands, innately ‘blind’ neurobiological receptors can be converted into photoreceptors, which allows them to be photoregulated with high spatiotemporal precision. Here, we present the optochemical control of neuronal nicotinic acetylcholine receptors (nAChRs) with photoswitchable tethered agonists and antagonists. Using structure-based design, we produced heteromeric α3β4 and α4β2 nAChRs that can be activated or inhibited with deep-violet light, but respond normally to acetylcholine in the dark. The generation of these engineered receptors should facilitate investigation of the physiological and pathological functions of neuronal nAChRs and open a general pathway to photosensitizing pentameric ligand-gated ion channels.

  9. Surveys suck: Consumer preferences when purchasing genetically engineered foods.

    Science.gov (United States)

    Powell, Douglas A

    2013-01-01

    Many studies have attempted to gauge consumers' acceptance of genetically engineered or modified (GM) foods. Surveys, asking people about attitudes and intentions, are easy-to-collect proxies of consumer behavior. However, participants tend to respond as citizens of society, not discrete individuals, thereby inaccurately portraying their potential behavior. The Theory of Planned Behavior improved the accuracy of self-reported information, but its limited capacity to account for intention variance has been attributed to the hypothetical scenarios to which survey participants must respond. Valuation methods, asking how much consumers may be willing to pay or accept for GM foods, have revealed that consumers are usually willing to accept them at some price, or in some cases willing to pay a premium. Ultimately, it's consumers' actual--not intended--behavior that is of most interest to policy makers and business decision-makers. Real choice experiments offer the best avenue for revealing consumers' food choices in normal life.

  10. [Histocompatibility of nano-hydroxyapatite/poly-co-glycolic acid tissue engineering bone modified by mesenchymal stem cells with vascular endothelial frowth factor].

    Science.gov (United States)

    Zhang, Minglei; Wang, Dapeng; Yin, Ruofeng

    2015-10-06

    To explorec Histocompatibility of nano-hydroxyapatite/poly-co-glycolic acid tissue engineering bone modified by mesenchymal stem cells with vascular endothelial frowth factor transinfected. Rat bone marrow mesenchymal stem cells (BMSCs) was separated, using BMSCs as target cells, and then vascular endothelial growth factor (VEGF) gene was transfected. Composite bone marrow mesenchymal stem cells and cells transfected with nano-hydroxyapatite (HA)/polylactic-co-glycolic acid (PLGA). The composition of cell and scaffold was observed. The blank plasmid transfection was 39.1%, 40.1% in VEGF group. The cell adhesion and growth was found on the scaffold pore wall after 5 days, and the number of adherent cells in the nano-HA/PLGA composite scaffold material basically had no significant difference in both. Although the nano-HA/PLGA scaffold material is still not fully meet the requirements of the matrix material for bone tissue engineering, but good biocompatibility, structure is its rich microporous satisfaction in material mechanics, toughening, enhanced obviously. Composition scaffold with BMSCs transfected by VEGF plasmid, the ability of angiogenesis is promoted.

  11. Genetic engineering of stem cells for enhanced therapy.

    Science.gov (United States)

    Nowakowski, Adam; Andrzejewska, Anna; Janowski, Miroslaw; Walczak, Piotr; Lukomska, Barbara

    2013-01-01

    Stem cell therapy is a promising strategy for overcoming the limitations of current treatment methods. The modification of stem cell properties may be necessary to fully exploit their potential. Genetic engineering, with an abundance of methodology to induce gene expression in a precise and well-controllable manner, is particularly attractive for this purpose. There are virus-based and non-viral methods of genetic manipulation. Genome-integrating viral vectors are usually characterized by highly efficient and long-term transgene expression, at a cost of safety. Non-integrating viruses are also highly efficient in transduction, and, while safer, offer only a limited duration of transgene expression. There is a great diversity of transfectable forms of nucleic acids; however, for efficient shuttling across cell membranes, additional manipulation is required. Both physical and chemical methods have been employed for this purpose. Stem cell engineering for clinical applications is still in its infancy and requires further research. There are two main strategies for inducing transgene expression in therapeutic cells: transient and permanent expression. In many cases, including stem cell trafficking and using cell therapy for the treatment of rapid-onset disease with a short healing process, transient transgene expression may be a sufficient and optimal approach. For that purpose, mRNA-based methods seem ideally suited, as they are characterized by a rapid, highly efficient transfection, with outstanding safety. Permanent transgene expression is primarily based on the application of viral vectors, and, due to safety concerns, these methods are more challenging. There is active, ongoing research toward the development of non-viral methods that would induce permanent expression, such as transposons and mammalian artificial chromosomes.

  12. Entomic Resistance Genes for Genetic Engineering in Agricultural Furtherance

    Directory of Open Access Journals (Sweden)

    Pankaj Kumar

    2015-02-01

    Full Text Available Genetic engineering for insect pest’s management in crop plants offers the potential of a user-friendly, environmentfriendly and consumer-friendly method of crop protection to meet the demands of sustainable agriculture. Food and energy insecurities are currently two foremost problems being faced worldwide. Losses due to pests and diseases have been estimated to be around 37% of the agricultural production worldwide, with 13% due to insects. Engineering insect resistance in transgenic plants has been achieved through the use of insect control protein genes of Bacillus thuringiensis. Till now, researchers have focused on the introduction of genes for expression of modified Bacillus thuringiensis (Bt toxins. Successful results on the control of Bt-susceptible pests have been achieved in the laboratory and finally in the field and now commercialized Bt transgenic crops are used worldwide. Other alternative methods exploit plant-derived insect control genes with promising results. Today insect-resistance transgenes, whether of plant, bacterial or other origin, can be introduced in to plants to increase the level of insect resistance so as to contribute to sustainable agricultural practices.

  13. Genetic engineering of platelets to neutralize circulating tumor cells.

    Science.gov (United States)

    Li, Jiahe; Sharkey, Charles C; Wun, Brittany; Liesveld, Jane L; King, Michael R

    2016-04-28

    Mounting experimental evidence demonstrates that platelets support cancer metastasis. Within the circulatory system, platelets guard circulating tumor cells (CTCs) from immune elimination and promote their arrest at the endothelium, supporting CTC extravasation into secondary sites. Neutralization of CTCs in blood circulation can potentially attenuate metastases to distant organs. Therefore, extensive studies have explored the blockade of platelet-CTC interactions as an anti-metastatic strategy. Such an intervention approach, however, may cause bleeding disorders since the platelet-CTC interactions inherently rely on the blood coagulation cascade including platelet activation. On the other hand, platelets have been genetically engineered to correct inherited bleeding disorders in both animal models and human clinical trials. In this study, inspired by the physical association between platelets and CTCs, platelets were genetically modified to express surface-bound tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), a cytokine known to induce apoptosis specifically in tumor cells. The TRAIL-expressing platelets were demonstrated to kill cancer cells in vitro and significantly reduce metastases in a mouse model of prostate cancer metastasis. Our results suggest that using platelets to produce and deliver cancer-specific therapeutics can provide a Trojan-horse strategy of neutralizing CTCs to attenuate metastasis.

  14. Genetic engineering and therapy for inherited and acquired cardiomyopathies.

    Science.gov (United States)

    Day, Sharlene; Davis, Jennifer; Westfall, Margaret; Metzger, Joseph

    2006-10-01

    The cardiac myofilaments consist of a highly ordered assembly of proteins that collectively generate force in a calcium-dependent manner. Defects in myofilament function and its regulation have been implicated in various forms of acquired and inherited human heart disease. For example, during cardiac ischemia, cardiac myocyte contractile performance is dramatically downregulated due in part to a reduced sensitivity of the myofilaments to calcium under acidic pH conditions. Over the last several years, the thin filament regulatory protein, troponin I, has been identified as an important mediator of this response. Mutations in troponin I and other sarcomere genes are also linked to several distinct inherited cardiomyopathic phenotypes, including hypertrophic, dilated, and restrictive cardiomyopathies. With the cardiac sarcomere emerging as a central player for such a diverse array of human heart diseases, genetic-based strategies that target the myofilament will likely have broad therapeutic potential. The development of safe vector systems for efficient gene delivery will be a critical hurdle to overcome before these types of therapies can be successfully applied. Nonetheless, studies focusing on the principles of acute genetic engineering of the sarcomere hold value as they lay the essential foundation on which to build potential gene-based therapies for heart disease.

  15. Increased production of nutriments by genetically engineered crops.

    Science.gov (United States)

    Sévenier, Robert; van der Meer, Ingrid M; Bino, Raoul; Koops, Andries J

    2002-06-01

    Plants are the basis of human nutrition and have been selected and improved to assure this purpose. Nowadays, new technologies such as genetic engineering and genomics approaches allow further improvement of plants. We describe here three examples for which these techniques have been employed. We introduced the first enzyme involved in fructan synthesis, the sucrose sucrose fructosyltransferase (isolated from Jerusalem artichoke), into sugar beet. The transgenic sugar beet showed a dramatic change in the nature of the accumulated sugar, 90% of the sucrose being converted into fructan. The use of transgenic sugar beet for the production and isolation of fructans will result in a more efficient plant production system of fructans and should promote their use in human food. The second example shows how the over-expression of the key enzyme of flavonoid biosynthesis could increase anti-oxidant levels in tomato. Introduction of a highly expressed chalcone isomerase led to a seventyfold increase of the amount of quercetin glucoside, which is a strong anti-oxidant in tomato. We were also able to modify the essential amino acid content of potato in order to increase its nutritional value. The introduction of a feedback insensitive bacterial gene involved in biosynthesis of aspartate family amino acids led to a sixfold increase of the lysine content. Because the use of a bacterial gene could appear to be controversial, we also introduced a mutated form of the plant key enzyme of lysine biosynthesis (dihydrodipicolinate synthase) in potato. This modification led to a 15 times increase of the lysine content of potato. This increase of the essential amino acid lysine influences the nutritional value of potato, which normally has low levels of several essential amino acids. These three examples show how the metabolism of primary constituents of the plant cell such as sugar or amino acids, but also of secondary metabolites such as flavonoids, can be modified by genetic

  16. Genetic Engineering: A Matter that Requires Further Refinement in Spanish Secondary School Textbooks

    Science.gov (United States)

    Martinez-Gracia, M. V.; Gil-Quylez, M. J.; Osada, J.

    2003-01-01

    Genetic engineering is now an integral part of many high school textbooks but little work has been done to assess whether it is being properly addressed. A checklist with 19 items was used to analyze how genetic engineering is presented in biology textbooks commonly used in Spanish high schools, including the content, its relationship with…

  17. 78 FR 13302 - Syngenta Biotechnology, Inc.; Determination of Nonregulated Status of Corn Genetically Engineered...

    Science.gov (United States)

    2013-02-27

    ... Products Altered or Produced Through Genetic Engineering Which Are Plant Pests or Which There Is Reason to... movement, or release into the environment) of organisms and products altered or produced through genetic engineering that are plant pests or that there is reason to believe are plant pests. Such...

  18. Long term culture of mesenchymal stem cells in hypoxia promotes a genetic program maintaining their undifferentiated and multipotent status

    Directory of Open Access Journals (Sweden)

    de Carvalho Marcelo

    2011-03-01

    Full Text Available Abstract Background In the bone marrow, hematopietic and mesenchymal stem cells form a unique niche in which the oxygen tension is low. Hypoxia may have a role in maintaining stem cell fate, self renewal and multipotency. However, whereas most studies addressed the effect of transient in vitro exposure of MSC to hypoxia, permanent culture under hypoxia should reflect the better physiological conditions. Results Morphologic studies, differentiation and transcriptional profiling experiments were performed on MSC cultured in normoxia (21% O2 versus hypoxia (5% O2 for up to passage 2. Cells at passage 0 and at passage 2 were compared, and those at passage 0 in hypoxia generated fewer and smaller colonies than in normoxia. In parallel, MSC displayed (>4 fold inhibition of genes involved in DNA metabolism, cell cycle progression and chromosome cohesion whereas transcripts involved in adhesion and metabolism (CD93, ESAM, VWF, PLVAP, ANGPT2, LEP, TCF1 were stimulated. Compared to normoxic cells, hypoxic cells were morphologically undifferentiated and contained less mitochondrias. After this lag phase, cells at passage 2 in hypoxia outgrew the cells cultured in normoxia and displayed an enhanced expression of genes (4-60 fold involved in extracellular matrix assembly (SMOC2, neural and muscle development (NOG, GPR56, SNTG2, LAMA and epithelial development (DMKN. This group described herein for the first time was assigned by the Gene Ontology program to "plasticity". Conclusion The duration of hypoxemia is a critical parameter in the differentiation capacity of MSC. Even in growth promoting conditions, hypoxia enhanced a genetic program that maintained the cells undifferentiated and multipotent. This condition may better reflect the in vivo gene signature of MSC, with potential implications in regenerative medicine.

  19. Non-genetic engineering of cells for drug delivery and cell-based therapy.

    Science.gov (United States)

    Wang, Qun; Cheng, Hao; Peng, Haisheng; Zhou, Hao; Li, Peter Y; Langer, Robert

    2015-08-30

    Cell-based therapy is a promising modality to address many unmet medical needs. In addition to genetic engineering, material-based, biochemical, and physical science-based approaches have emerged as novel approaches to modify cells. Non-genetic engineering of cells has been applied in delivering therapeutics to tissues, homing of cells to the bone marrow or inflammatory tissues, cancer imaging, immunotherapy, and remotely controlling cellular functions. This new strategy has unique advantages in disease therapy and is complementary to existing gene-based cell engineering approaches. A better understanding of cellular systems and different engineering methods will allow us to better exploit engineered cells in biomedicine. Here, we review non-genetic cell engineering techniques and applications of engineered cells, discuss the pros and cons of different methods, and provide our perspectives on future research directions. Copyright © 2014 Elsevier B.V. All rights reserved.

  20. Mesenchymal stem cells and myoblast differentiation under HGF and IGF-1 stimulation for 3D skeletal muscle tissue engineering.

    Science.gov (United States)

    Witt, R; Weigand, A; Boos, A M; Cai, A; Dippold, D; Boccaccini, A R; Schubert, D W; Hardt, M; Lange, C; Arkudas, A; Horch, R E; Beier, J P

    2017-02-28

    Volumetric muscle loss caused by trauma or after tumour surgery exceeds the natural regeneration capacity of skeletal muscle. Hence, the future goal of tissue engineering (TE) is the replacement and repair of lost muscle tissue by newly generating skeletal muscle combining different cell sources, such as myoblasts and mesenchymal stem cells (MSCs), within a three-dimensional matrix. Latest research showed that seeding skeletal muscle cells on aligned constructs enhance the formation of myotubes as well as cell alignment and may provide a further step towards the clinical application of engineered skeletal muscle. In this study the myogenic differentiation potential of MSCs upon co-cultivation with myoblasts and under stimulation with hepatocyte growth factor (HGF) and insulin-like growth factor-1 (IGF-1) was evaluated. We further analysed the behaviour of MSC-myoblast co-cultures in different 3D matrices. Primary rat myoblasts and rat MSCs were mono- and co-cultivated for 2, 7 or 14 days. The effect of different concentrations of HGF and IGF-1 alone, as well as in combination, on myogenic differentiation was analysed using microscopy, multicolour flow cytometry and real-time PCR. Furthermore, the influence of different three-dimensional culture models, such as fibrin, fibrin-collagen-I gels and parallel aligned electrospun poly-ε-caprolacton collagen-I nanofibers, on myogenic differentiation was analysed. MSCs could be successfully differentiated into the myogenic lineage both in mono- and in co-cultures independent of HGF and IGF-1 stimulation by expressing desmin, myocyte enhancer factor 2, myosin heavy chain 2 and alpha-sarcomeric actinin. An increased expression of different myogenic key markers could be observed under HGF and IGF-1 stimulation. Even though, stimulation with HGF/IGF-1 does not seem essential for sufficient myogenic differentiation. Three-dimensional cultivation in fibrin-collagen-I gels induced higher levels of myogenic differentiation

  1. Development of VEGF-loaded PLGA matrices in association with mesenchymal stem cells for tissue engineering.

    Science.gov (United States)

    Rosa, A R; Steffens, D; Santi, B; Quintiliano, K; Steffen, N; Pilger, D A; Pranke, P

    2017-08-07

    The association of bioactive molecules, such as vascular endothelial growth factor (VEGF), with nanofibers facilitates their controlled release, which could contribute to cellular migration and differentiation in tissue regeneration. In this research, the influence of their incorporation on a polylactic-co-glycolic acid (PLGA) scaffold produced by electrospinning on cell adhesion and viability and cytotoxicity was carried out in three groups: 1) PLGA/BSA/VEGF; 2) PLGA/BSA, and 3) PLGA. Morphology, fiber diameter, contact angle, loading efficiency and controlled release of VEGF of the biomaterials, among others, were measured. The nanofibers showed smooth surfaces without beads and with interconnected pores. PLGA/BSA/VEGF showed the smallest water contact angle and VEGF released for up to 160 h. An improvement in cell adhesion was observed for the PLGA/BSA/VEGF scaffolds compared to the other groups and the scaffolds were non-toxic for the cells. Therefore, the scaffolds were shown to be a good strategy for sustained delivery of VEGF and may be a useful tool for tissue engineering.

  2. Development of VEGF-loaded PLGA matrices in association with mesenchymal stem cells for tissue engineering

    Directory of Open Access Journals (Sweden)

    A.R. Rosa

    Full Text Available The association of bioactive molecules, such as vascular endothelial growth factor (VEGF, with nanofibers facilitates their controlled release, which could contribute to cellular migration and differentiation in tissue regeneration. In this research, the influence of their incorporation on a polylactic-co-glycolic acid (PLGA scaffold produced by electrospinning on cell adhesion and viability and cytotoxicity was carried out in three groups: 1 PLGA/BSA/VEGF; 2 PLGA/BSA, and 3 PLGA. Morphology, fiber diameter, contact angle, loading efficiency and controlled release of VEGF of the biomaterials, among others, were measured. The nanofibers showed smooth surfaces without beads and with interconnected pores. PLGA/BSA/VEGF showed the smallest water contact angle and VEGF released for up to 160 h. An improvement in cell adhesion was observed for the PLGA/BSA/VEGF scaffolds compared to the other groups and the scaffolds were non-toxic for the cells. Therefore, the scaffolds were shown to be a good strategy for sustained delivery of VEGF and may be a useful tool for tissue engineering.

  3. A FIELD STUDY WITH GENETICALLY ENGINEERED ALFALFA INOCULATED WITH RECOMBINANT SINORHIZOBIUM MELILOTI: EFFECTS ON THE SOIL ECOSYSTEM

    Science.gov (United States)

    The agricultural use of genetically engineered plants and microorganisms has become increasingly common. Because genetically engineered plants and microorganisms can produce compounds foreign to their environment, there is concern that they may become established outside of thei...

  4. A FIELD STUDY WITH GENETICALLY ENGINEERED ALFALFA INOCULATED WITH RECOMBINANT SINORHIZOBIUM MELILOTI: EFFECTS ON THE SOIL ECOSYSTEM

    Science.gov (United States)

    The agricultural use of genetically engineered plants and microorganisms has become increasingly common. Because genetically engineered plants and microorganisms can produce compounds foreign to their environment, there is concern that they may become established outside of thei...

  5. Genetically engineered microorganisms for the detection of explosives' residues

    Directory of Open Access Journals (Sweden)

    Benjamin eShemer

    2015-10-01

    Full Text Available The manufacture and use of explosives throughout the past century has resulted in the extensive pollution of soils and groundwater, and the widespread interment of landmines imposes a major humanitarian risk and prevents civil development of large areas. As most current landmine detection technologies require actual presence at the surveyed areas, thus posing a significant risk to personnel, diverse research efforts are aimed at the development of remote detection solutions. One possible means proposed to fulfill this objective is the use of microbial bioreporters: genetically engineered microorganisms tailored to generate an optical signal in the presence of explosives’ vapors. The use of such sensor bacteria will allow to pinpoint the locations of explosive devices in a minefield. While no study has yet resulted in a commercially operational system, significant progress has been made in the design and construction of explosives-sensing bacterial strains. In this article we review the attempts to construct microbial bioreporters for the detection of explosives, and analyze the steps that need to be undertaken for this strategy to be applicable for landmine detection.

  6. Genetically engineered microorganisms for the detection of explosives’ residues

    Science.gov (United States)

    Shemer, Benjamin; Palevsky, Noa; Yagur-Kroll, Sharon; Belkin, Shimshon

    2015-01-01

    The manufacture and use of explosives throughout the past century has resulted in the extensive pollution of soils and groundwater, and the widespread interment of landmines imposes a major humanitarian risk and prevents civil development of large areas. As most current landmine detection technologies require actual presence at the surveyed areas, thus posing a significant risk to personnel, diverse research efforts are aimed at the development of remote detection solutions. One possible means proposed to fulfill this objective is the use of microbial bioreporters: genetically engineered microorganisms “tailored” to generate an optical signal in the presence of explosives’ vapors. The use of such sensor bacteria will allow to pinpoint the locations of explosive devices in a minefield. While no study has yet resulted in a commercially operational system, significant progress has been made in the design and construction of explosives-sensing bacterial strains. In this article we review the attempts to construct microbial bioreporters for the detection of explosives, and analyze the steps that need to be undertaken for this strategy to be applicable for landmine detection. PMID:26579085

  7. Genetic engineering in Cowpea (Vigna unguiculata): history, status and prospects.

    Science.gov (United States)

    Citadin, Cristiane T; Ibrahim, Abdulrazak B; Aragão, Francisco J L

    2011-01-01

    In the last three decades, a number of attempts have been made to develop reproducible protocols for generating transgenic cowpea that permit the expression of genes of agronomic importance. Pioneer works focused on the development of such systems vis-à-vis an in vitro culture system that would guarantee de novo regeneration of transgenic cowpea arising from cells amenable to one form of gene delivery system or another, but any such system has eluded researchers over the years. Despite this apparent failure, significant progress has been made in generating transgenic cowpea, bringing researchers much nearer to their goal than thirty years ago. Now, various researchers have successfully established transgenic procedures for cowpea with evidence of inherent transgenes of interest, effected by progenies in a Mendelian fashion. New opportunities have thus emerged to optimize existing protocols and devise new strategies to ensure the development of transgenic cowpea with desirable agronomic traits. This review chronicles the important milestones in the last thirty years that have marked the evolution of genetic engineering of cowpea. It also highlights the progress made and describes new strategies that have arisen, culminating in the current status of transgenic technologies for cowpea.

  8. Novel pancreatic cancer cell lines derived from genetically engineered mouse models of spontaneous pancreatic adenocarcinoma: applications in diagnosis and therapy.

    Directory of Open Access Journals (Sweden)

    María P Torres

    Full Text Available Pancreatic cancer (PC remains one of the most lethal human malignancies with poor prognosis. Despite all advances in preclinical research, there have not been significant translation of novel therapies into the clinics. The development of genetically engineered mouse (GEM models that produce spontaneous pancreatic adenocarcinoma (PDAC have increased our understanding of the pathogenesis of the disease. Although these PDAC mouse models are ideal for studying potential therapies and specific genetic mutations, there is a need for developing syngeneic cell lines from these models. In this study, we describe the successful establishment and characterization of three cell lines derived from two (PDAC mouse models. The cell line UN-KC-6141 was derived from a pancreatic tumor of a Kras(G12D;Pdx1-Cre (KC mouse at 50 weeks of age, whereas UN-KPC-960 and UN-KPC-961 cell lines were derived from pancreatic tumors of Kras(G12D;Trp53(R172H;Pdx1-Cre (KPC mice at 17 weeks of age. The cancer mutations of these parent mice carried over to the daughter cell lines (i.e. Kras(G12D mutation was observed in all three cell lines while Trp53 mutation was observed only in KPC cell lines. The cell lines showed typical cobblestone epithelial morphology in culture, and unlike the previously established mouse PDAC cell line Panc02, expressed the ductal marker CK19. Furthermore, these cell lines expressed the epithelial-mesenchymal markers E-cadherin and N-cadherin, and also, Muc1 and Muc4 mucins. In addition, these cell lines were resistant to the chemotherapeutic drug Gemcitabine. Their implantation in vivo produced subcutaneous as well as tumors in the pancreas (orthotopic. The genetic mutations in these cell lines mimic the genetic compendium of human PDAC, which make them valuable models with a high potential of translational relevance for examining diagnostic markers and therapeutic drugs.

  9. Induction of Atherosclerosis in Mice and Hamsters Without Germline Genetic Engineering

    DEFF Research Database (Denmark)

    Bjørklund, Martin Mæng; Hollensen, Anne K; Hagensen, Mette K

    2014-01-01

    RATIONALE: Atherosclerosis can be achieved in animals by germline genetic engineering, leading to hypercholesterolemia, but such models are constrained to few species and strains, and they are difficult to combine with other powerful techniques involving genetic manipulation or variation. OBJECTIVE......: To develop a method for induction of atherosclerosis without germline genetic engineering. METHODS AND RESULTS: Recombinant adeno-associated viral vectors were engineered to encode gain-of-function proprotein convertase subtilisin/kexin type 9 mutants, and mice were given a single intravenous vector...... are a rapid and versatile method to induce atherosclerosis in animals. This method should prove useful for experiments that are high-throughput or involve genetic techniques, strains, or species that do not combine well with current genetically engineered models....

  10. Developing Novel Therapeutic Approaches in Small Cell Lung Carcinoma Using Genetically Engineered Mouse Models and Human Circulating Tumor Cells

    Science.gov (United States)

    2015-10-01

    Using Genetically Engineered Mouse Models and Human Circulating Tumor Cells PRINCIPAL INVESTIGATOR: Jeffrey Engelman MD PhD CONTRACTING...SUBTITLE Developiing Novel Therapeutic Approaches in Small Cell Lung 5a. CONTRACT NUMBER Carcinoma Using Genetically Engineered Mouse Models and 5b...biomarkers. 15. SUBJECT TERMS Small cell lung cancer (SCLC), Genetically engineered mouse model (GEMM), BH3 mimetic, TORC inhibitor, Apoptosis

  11. Effect of a genetically engineered bacteriophage on Enterococcus faecalis biofilms.

    Science.gov (United States)

    Tinoco, Justine Monnerat; Buttaro, Bettina; Zhang, Hongming; Liss, Nadia; Sassone, Luciana; Stevens, Roy

    2016-11-01

    Enterococcus faecalis is a Gram-positive, facultative anaerobic bacterium that is associated with failed endodontic cases and nosocomial infections. E. faecalis can form biofilms, penetrate dentinal tubules and survive in root canals with scarce nutritional supplies. These properties can make E. faecalis resistant to conventional endodontic disinfection therapy. Furthermore, treatment may be complicated by the fact that many E. faecalis strains are resistant to antibiotics. A potential alternative to antibiotic therapy is phage therapy. ϕEf11 is a temperate phage that infects strains of E. faecalis. It was previously sequenced and genetically engineered to modify its properties in order to render it useful as a therapeutic agent in phage therapy. In the current study, we have further genetically modified the phage to create phage ϕEf11/ϕFL1C(Δ36)(PnisA). The aim of this study was to evaluate the efficacy of bacteriophage ϕEf11/ϕFL1C(Δ36)(PnisA), to disrupt biofilms of two Enterococcus faecalis strains: JH2-2 (vancomycin-sensitive) and V583 (vancomycin-resistant). 24h static biofilms of E. faecalis strains JH2-2(pMSP3535 nisR/K) and V583 (pMSP3535nisR/K), formed on cover slips, were inoculated with bacteriophage ϕEf11/ϕFL1C(Δ36)(PnisA). After 24 and 48h incubation, the bacterial biomass was imaged by confocal microscopy and viable cells were quantified by colony forming unit measurement. The results showed a 10-100-fold decrease in viable cells (CFU/biofilm) after phage treatment, which was consistent with comparisons of treated and untreated biofilm images visualized as max projections of the Z-series. The biomass of both vancomycin-sensitive and vancomycin-resistant E. faecalis biofilms is markedly reduced following infection by bacteriophage ϕEf11/ϕFL1C(Δ36)(PnisA). Copyright © 2016 Elsevier Ltd. All rights reserved.

  12. In Vitro Testing of Scaffolds for Mesenchymal Stem Cell-Based Meniscus Tissue Engineering-Introducing a New Biocompatibility Scoring System.

    Science.gov (United States)

    Achatz, Felix P; Kujat, Richard; Pfeifer, Christian G; Koch, Matthias; Nerlich, Michael; Angele, Peter; Zellner, Johannes

    2016-04-07

    A combination of mesenchymal stem cells (MSCs) and scaffolds seems to be a promising approach for meniscus repair. To facilitate the search for an appropriate scaffold material a reliable and objective in vitro testing system is essential. This paper introduces a new scoring for this purpose and analyzes a hyaluronic acid (HA) gelatin composite scaffold and a polyurethane scaffold in combination with MSCs for tissue engineering of meniscus. The pore quality and interconnectivity of pores of a HA gelatin composite scaffold and a polyurethane scaffold were analyzed by surface photography and Berliner-Blau-BSA-solution vacuum filling. Further the two scaffold materials were vacuum-filled with human MSCs and analyzed by histology and immunohistochemistry after 21 days in chondrogenic media to determine cell distribution and cell survival as well as proteoglycan production, collagen type I and II content. The polyurethane scaffold showed better results than the hyaluronic acid gelatin composite scaffold, with signs of central necrosis in the HA gelatin composite scaffolds. The polyurethane scaffold showed good porosity, excellent pore interconnectivity, good cell distribution and cell survival, as well as an extensive content of proteoglycans and collagen type II. The polyurethane scaffold seems to be a promising biomaterial for a mesenchymal stem cell-based tissue engineering approach for meniscal repair. The new score could be applied as a new standard for in vitro scaffold testing.

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

    Science.gov (United States)

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

    1997-09-01

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

  14. Laboratory injection molder for the fabrication of polymeric porous poly-epsilon-caprolactone scaffolds for preliminary mesenchymal stem cells tissue engineering applications

    KAUST Repository

    Limongi, Tania

    2016-12-16

    This study presents a simple and rapid fabrication technique involving injection molding and particle leaching (IM/PL) to fabricate the porous scaffold for tissue engineering applications. Sodium chloride (NaCl) and Sucrose are separately mixed with the poly-epsilon-caprolactone (PCL) granules using a screwed thermo regulated extruder, than the biocompatible scaffolds are fabricated through injection molding. The micro/nano structure of the samples and their different grade of porosity were characterized by scanning electron microscopy and mercury intrusion porosimetry. Bone marrow-derived mesenchymal stem cells are chose to cell culture and Hoechst 33342 staining was used to verify the biocompatibility of the polymeric porous surfaces. We concluded that, by using the same fast solvent free injection/leaching process, the use of Sucrose as porogen, instead of NaCl, allowed the obtainment of biocompatible scaffolds with a higher grade of porosity with suitable cell adhesion capacity for tissue engineering purpose.

  15. Molecular profiling techniques as tools to detect potential unintended effects in genetically engineered maize

    CSIR Research Space (South Africa)

    Barros, E

    2010-05-01

    Full Text Available In the early stages of production and commercialization of foods derived from genetically engineered (GE) plants, international consensus was reached on the principles of food safety evaluation. The concept of substantial equivalence became...

  16. Gene flow in genetically engineered perennial grasses: Lessons for modification of dedicated bioenergy crops

    Science.gov (United States)

    The potential ecological consequences of the commercialization of genetically engineered (GD) crops have been the subject of intense debate, particularly when the GE crops are perennial and capable of outcrossing to wild relatives. The essential ecological impact issues for engi...

  17. IMPROVING PLANT GENETIC ENGINEERING BY MANIPULATING THE HOST. (R829479C001)

    Science.gov (United States)

    Agrobacterium-mediated transformation is a major technique for the genetic engineering of plants. However, there are many economically important crop and tree species that remain highly recalcitrant to Agrobacterium infection. Although attempts have been made to ...

  18. Genetic engineering of human ES and iPS cells using TALE nucleases

    OpenAIRE

    Hockemeyer, Dirk; Wang, Haoyi; Kiani, Samira; Lai, Christine S.; Gao, Qing; Cassady, John P.; Cost, Gregory J.; Zhang, Lei; Santiago, Yolanda; Miller, Jeffrey C; Zeitler, Bryan; Cherone, Jennifer M.; Meng, Xiangdong; Hinkley, Sarah J; Rebar, Edward J.

    2011-01-01

    Targeted genetic engineering of human pluripotent cells is a prerequisite for exploiting their full potential. Such genetic manipulations can be achieved using site-specific nucleases. Here we engineered transcription activator–like effector nucleases (TALENs) for five distinct genomic loci. At all loci tested we obtained human embryonic stem cell (ESC) and induced pluripotent stem cell (iPSC) clones carrying transgenic cassettes solely at the TALEN-specified location. Our data suggest that T...

  19. Genetically engineered cardiac pacemaker: Stem cells transfected with HCN2 gene and myocytes-A model

    Energy Technology Data Exchange (ETDEWEB)

    Kanani, S. [Institut Genomique Fonctionelle, 141 Rue de la Cardonille, 34396 Montpellier (France); Institut Non Lineaire de Nice, CNRS and Universite de Nice, 1361 route des Lucioles, 06560 Valbonne (France); Pumir, A. [Institut Non Lineaire de Nice, CNRS and Universite de Nice, 1361 route des Lucioles, 06560 Valbonne (France); Laboratoire J.A. Dieudonne, CNRS and Universite de Nice, Parc Valrose, 06108 Nice (France)], E-mail: alain.pumir@unice.fr; Krinsky, V. [Institut Non Lineaire de Nice, CNRS and Universite de Nice, 1361 route des Lucioles, 06560 Valbonne (France)

    2008-01-07

    One of the successfully tested methods to design genetically engineered cardiac pacemaker cells consists in transfecting a human mesenchymal stem cell (hMSC) with a HCN2 gene and connecting it to a myocyte. We develop and study a mathematical model, describing a myocyte connected to a hMSC transfected with a HCN2 gene. The cardiac action potential is described both with the simple Beeler-Reuter model, as well as with the elaborate dynamic Luo-Rudy model. The HCN2 channel is described by fitting electrophysiological records, in the spirit of Hodgkin-Huxley. The model shows that oscillations can occur in a pair myocyte-stem cell, that was not observed in the experiments yet. The model predicted that: (1) HCN pacemaker channels can induce oscillations only if the number of expressed I{sub K1} channels is low enough. At too high an expression level of I{sub K1} channels, oscillations cannot be induced, no matter how many pacemaker channels are expressed. (2) At low expression levels of I{sub K1} channels, a large domain of values in the parameter space (n, N) exists, where oscillations should be observed. We denote N the number of expressed pacemaker channels in the stem cell, and n the number of gap junction channels coupling the stem cell and the myocyte. (3) The expression levels of I{sub K1} channels observed in ventricular myocytes, both in the Beeler-Reuter and in the dynamic Luo-Rudy models are too high to allow to observe oscillations. With expression levels below {approx}1/4 of the original value, oscillations can be observed. The main consequence of this work is that in order to obtain oscillations in an experiment with a myocyte-stem cell pair, increasing the values of n, N is unlikely to be helpful, unless the expression level of I{sub K1} has been reduced enough. The model also allows us to explore levels of gene expression not yet achieved in experiments, and could be useful to plan new experiments, aimed at improving the robustness of the oscillations.

  20. Genetically engineered cardiac pacemaker: Stem cells transfected with HCN2 gene and myocytes—A model

    Science.gov (United States)

    Kanani, S.; Pumir, A.; Krinsky, V.

    2008-01-01

    One of the successfully tested methods to design genetically engineered cardiac pacemaker cells consists in transfecting a human mesenchymal stem cell (hMSC) with a HCN2 gene and connecting it to a myocyte. We develop and study a mathematical model, describing a myocyte connected to a hMSC transfected with a HCN2 gene. The cardiac action potential is described both with the simple Beeler Reuter model, as well as with the elaborate dynamic Luo Rudy model. The HCN2 channel is described by fitting electrophysiological records, in the spirit of Hodgkin Huxley. The model shows that oscillations can occur in a pair myocyte-stem cell, that was not observed in the experiments yet. The model predicted that: (1) HCN pacemaker channels can induce oscillations only if the number of expressed I channels is low enough. At too high an expression level of I channels, oscillations cannot be induced, no matter how many pacemaker channels are expressed. (2) At low expression levels of I channels, a large domain of values in the parameter space (n, N) exists, where oscillations should be observed. We denote N the number of expressed pacemaker channels in the stem cell, and n the number of gap junction channels coupling the stem cell and the myocyte. (3) The expression levels of I channels observed in ventricular myocytes, both in the Beeler Reuter and in the dynamic Luo Rudy models are too high to allow to observe oscillations. With expression levels below ˜1/4 of the original value, oscillations can be observed. The main consequence of this work is that in order to obtain oscillations in an experiment with a myocyte-stem cell pair, increasing the values of n, N is unlikely to be helpful, unless the expression level of I has been reduced enough. The model also allows us to explore levels of gene expression not yet achieved in experiments, and could be useful to plan new experiments, aimed at improving the robustness of the oscillations.

  1. Genome-scale genetic engineering in Escherichia coli.

    Science.gov (United States)

    Jeong, Jaehwan; Cho, Namjin; Jung, Daehee; Bang, Duhee

    2013-11-01

    Genome engineering has been developed to create useful strains for biological studies and industrial uses. However, a continuous challenge remained in the field: technical limitations in high-throughput screening and precise manipulation of strains. Today, technical improvements have made genome engineering more rapid and efficient. This review introduces recent advances in genome engineering technologies applied to Escherichia coli as well as multiplex automated genome engineering (MAGE), a recent technique proposed as a powerful toolkit due to its straightforward process, rapid experimental procedures, and highly efficient properties.

  2. Generating Alternative Engineering Designs by Integrating Desktop VR with Genetic Algorithms

    Science.gov (United States)

    Chandramouli, Magesh; Bertoline, Gary; Connolly, Patrick

    2009-01-01

    This study proposes an innovative solution to the problem of multiobjective engineering design optimization by integrating desktop VR with genetic computing. Although, this study considers the case of construction design as an example to illustrate the framework, this method can very much be extended to other engineering design problems as well.…

  3. Generating Alternative Engineering Designs by Integrating Desktop VR with Genetic Algorithms

    Science.gov (United States)

    Chandramouli, Magesh; Bertoline, Gary; Connolly, Patrick

    2009-01-01

    This study proposes an innovative solution to the problem of multiobjective engineering design optimization by integrating desktop VR with genetic computing. Although, this study considers the case of construction design as an example to illustrate the framework, this method can very much be extended to other engineering design problems as well.…

  4. Teaching Applied Genetics and Molecular Biology to Agriculture Engineers. Application of the European Credit Transfer System

    Science.gov (United States)

    Weiss, J.; Egea-Cortines, M.

    2008-01-01

    We have been teaching applied molecular genetics to engineers and adapted the teaching methodology to the European Credit Transfer System. We teach core principles of genetics that are universal and form the conceptual basis of most molecular technologies. The course then teaches widely used techniques and finally shows how different techniques…

  5. Ray Wu, Cornell's acclaimed pioneer of genetic engineering and developer of insect-resistant rice

    Institute of Scientific and Technical Information of China (English)

    2009-01-01

    @@ ITHACA, N.Y. - Ray J. Wu, Comell University professor of molecular biology and genetics, who was widely recognized as one of the fathers of genetic engineering and who developed and sought to feed the world with a higher yielding rice that resists insects and drought, died of cardiac arrest in Ithaca, Feb. 10.

  6. Genetic engineering of Ganoderma lucidum for the efficient production of ganoderic acids.

    Science.gov (United States)

    Xu, Jun-Wei; Zhong, Jian-Jiang

    2015-01-01

    Ganoderma lucidum is a well-known traditional medicinal mushroom that produces ganoderic acids with numerous interesting bioactivities. Genetic engineering is an efficient approach to improve ganoderic acid biosynthesis. However, reliable genetic transformation methods and appropriate genetic manipulation strategies remain underdeveloped and thus should be enhanced. We previously established a homologous genetic transformation method for G. lucidum; we also applied the established method to perform the deregulated overexpression of a homologous 3-hydroxy-3-methyl-glutaryl coenzyme A reductase gene in G. lucidum. Engineered strains accumulated more ganoderic acids than wild-type strains. In this report, the genetic transformation systems of G. lucidum are described; current trends are also presented to improve ganoderic acid production through the genetic manipulation of G. lucidum.

  7. The Significance of Content Knowledge for Informal Reasoning regarding Socioscientific Issues: Applying Genetics Knowledge to Genetic Engineering Issues

    Science.gov (United States)

    Sadler, Troy D.; Zeidler, Dana L.

    2005-01-01

    This study focused on informal reasoning regarding socioscientific issues. It sought to explore how content knowledge influenced the negotiation and resolution of contentious and complex scenarios based on genetic engineering. Two hundred and sixty-nine students drawn from undergraduate natural science and nonnatural science courses completed a…

  8. Biotechnology, Genetic Engineering and Society. Monograph Series: III.

    Science.gov (United States)

    Kieffer, George H.

    New techniques have expanded the field of biotechnology and awarded scientists an unprecedented degree of control over the genetic constitutions of living things. The knowledge of DNA science is the basis for this burgeoning industry which may be a major force in human existence. Just as it is possible to move genetic material from one organism to…

  9. Harnessing biodiesel-producing microbes: from genetic engineering of lipase to metabolic engineering of fatty acid biosynthetic pathway.

    Science.gov (United States)

    Yan, Jinyong; Yan, Yunjun; Madzak, Catherine; Han, Bingnan

    2017-02-01

    Microbial production routes, notably whole-cell lipase-mediated biotransformation and fatty-acids-derived biosynthesis, offer new opportunities for synthesizing biodiesel. They compare favorably to immobilized lipase and chemically catalyzed processes. Genetically modified whole-cell lipase-mediated in vitro route, together with in vivo and ex vivo microbial biosynthesis routes, constitutes emerging and rapidly developing research areas for effective production of biodiesel. This review presents recent advances in customizing microorganisms for producing biodiesel, via genetic engineering of lipases and metabolic engineering (including system regulation) of fatty-acids-derived pathways. Microbial hosts used include Escherichia coli, Saccharomyces cerevisiae, Pichia pastoris and Aspergillus oryzae. These microbial cells can be genetically modified to produce lipases under different forms: intracellularly expressed, secreted or surface-displayed. They can be metabolically redesigned and systematically regulated to obtain balanced biodiesel-producing cells, as highlighted in this study. Such genetically or metabolically modified microbial cells can support not only in vitro biotransformation of various common oil feedstocks to biodiesel, but also de novo biosynthesis of biodiesel from glucose, glycerol or even cellulosic biomass. We believe that the genetically tractable oleaginous yeast Yarrowia lipolytica could be developed to an effective biodiesel-producing microbial cell factory. For this purpose, we propose several engineered pathways, based on lipase and wax ester synthase, in this promising oleaginous host.

  10. Using genetically engineered animal models in the postgenomic era to understand gene function in alcoholism.

    Science.gov (United States)

    Reilly, Matthew T; Harris, R Adron; Noronha, Antonio

    2012-01-01

    Over the last 50 years, researchers have made substantial progress in identifying genetic variations that underlie the complex phenotype of alcoholism. Not much is known, however, about how this genetic variation translates into altered biological function. Genetic animal models recapitulating specific characteristics of the human condition have helped elucidate gene function and the genetic basis of disease. In particular, major advances have come from the ability to manipulate genes through a variety of genetic technologies that provide an unprecedented capacity to determine gene function in the living organism and in alcohol-related behaviors. Even newer genetic-engineering technologies have given researchers the ability to control when and where a specific gene or mutation is activated or deleted, allowing investigators to narrow the role of the gene's function to circumscribed neural pathways and across development. These technologies are important for all areas of neuroscience, and several public and private initiatives are making a new generation of genetic-engineering tools available to the scientific community at large. Finally, high-throughput "next-generation sequencing" technologies are set to rapidly increase knowledge of the genome, epigenome, and transcriptome, which, combined with genetically engineered mouse mutants, will enhance insight into biological function. All of these resources will provide deeper insight into the genetic basis of alcoholism.

  11. Genetic Networks of Complex Disorders: from a Novel Search Engine for PubMed Article Database.

    Science.gov (United States)

    Jung, Jae-Yoon; Wall, Dennis Paul

    2013-01-01

    Finding genetic risk factors of complex disorders may involve reviewing hundreds of genes or thousands of research articles iteratively, but few tools have been available to facilitate this procedure. In this work, we built a novel publication search engine that can identify target-disorder specific, genetics-oriented research articles and extract the genes with significant results. Preliminary test results showed that the output of this engine has better coverage in terms of genes or publications, than other existing applications. We consider it as an essential tool for understanding genetic networks of complex disorders.

  12. Tools for genetic engineering of the yeast Hansenula polymorpha

    NARCIS (Netherlands)

    Saraya, Ruchi; Gidijala, Loknath; Veenhuis, Marten; van der Klei, Ida J; Mapelli, Valeria

    2014-01-01

    Hansenula polymorpha is a methylotrophic yeast species that has favorable properties for heterologous protein production and metabolic engineering. It provides an attractive expression platform with the capability to secrete high levels of commercially important proteins. Over the past few years

  13. Genetically engineered plants with increased vegetative oil content

    Science.gov (United States)

    Benning, Christoph

    2017-05-23

    The invention relates to genetically modified agricultural plants with increased oil content in vegetative tissues, as well as to expression systems, plant cells, seeds and vegetative tissues related thereto.

  14. Hybrid Neural-Network: Genetic Algorithm Technique for Aircraft Engine Performance Diagnostics Developed and Demonstrated

    Science.gov (United States)

    Kobayashi, Takahisa; Simon, Donald L.

    2002-01-01

    As part of the NASA Aviation Safety Program, a unique model-based diagnostics method that employs neural networks and genetic algorithms for aircraft engine performance diagnostics has been developed and demonstrated at the NASA Glenn Research Center against a nonlinear gas turbine engine model. Neural networks are applied to estimate the internal health condition of the engine, and genetic algorithms are used for sensor fault detection, isolation, and quantification. This hybrid architecture combines the excellent nonlinear estimation capabilities of neural networks with the capability to rank the likelihood of various faults given a specific sensor suite signature. The method requires a significantly smaller data training set than a neural network approach alone does, and it performs the combined engine health monitoring objectives of performance diagnostics and sensor fault detection and isolation in the presence of nominal and degraded engine health conditions.

  15. BMP2 genetically engineered MSCs and EPCs promote vascularized bone regeneration in rat critical-sized calvarial bone defects.

    Directory of Open Access Journals (Sweden)

    Xiaoning He

    Full Text Available Current clinical therapies for critical-sized bone defects (CSBDs remain far from ideal. Previous studies have demonstrated that engineering bone tissue using mesenchymal stem cells (MSCs is feasible. However, this approach is not effective for CSBDs due to inadequate vascularization. In our previous study, we have developed an injectable and porous nano calcium sulfate/alginate (nCS/A scaffold and demonstrated that nCS/A composition is biocompatible and has proper biodegradability for bone regeneration. Here, we hypothesized that the combination of an injectable and porous nCS/A with bone morphogenetic protein 2 (BMP2 gene-modified MSCs and endothelial progenitor cells (EPCs could significantly enhance vascularized bone regeneration. Our results demonstrated that delivery of MSCs and EPCs with the injectable nCS/A scaffold did not affect cell viability. Moreover, co-culture of BMP2 gene-modified MSCs and EPCs dramatically increased osteoblast differentiation of MSCs and endothelial differentiation of EPCs in vitro. We further tested the multifunctional bone reconstruction system consisting of an injectable and porous nCS/A scaffold (mimicking the nano-calcium matrix of bone and BMP2 genetically-engineered MSCs and EPCs in a rat critical-sized (8 mm caviarial bone defect model. Our in vivo results showed that, compared to the groups of nCS/A, nCS/A+MSCs, nCS/A+MSCs+EPCs and nCS/A+BMP2 gene-modified MSCs, the combination of BMP2 gene -modified MSCs and EPCs in nCS/A dramatically increased the new bone and vascular formation. These results demonstrated that EPCs increase new vascular growth, and that BMP2 gene modification for MSCs and EPCs dramatically promotes bone regeneration. This system could ultimately enable clinicians to better reconstruct the craniofacial bone and avoid donor site morbidity for CSBDs.

  16. DECOMPOSTION OF GENETICALLY ENGINEERED TOBACCO UNDER FIELD CONDITIONS: PERSISTENCE OF THE PROTEINASE INHIBITOR I PRODUCT AND EFFECTS OF SOIL MICROBIAL RESPIRATION AND PROTOZOA, NEMATODE AND MICROARTHR

    Science.gov (United States)

    1. To evaluate the potential effects of genetically engineered (transgenic) plants on soil ecosystems, litterbags containing leaves of non-engineered (parental) and transgenic tobacco plants were buried in field plots. The transgenic tobacco plants were genetically engineered to ...

  17. The use of genetically-engineered animals in science: perspectives of Canadian Animal Care Committee members.

    Science.gov (United States)

    Ormandy, Elisabeth H; Dale, Julie; Griffin, Gilly

    2013-05-01

    The genetic engineering of animals for their use in science challenges the implementation of refinement and reduction in several areas, including the invasiveness of the procedures involved, unanticipated welfare concerns, and the numbers of animals required. Additionally, the creation of genetically-engineered animals raises problems with the Canadian system of reporting animal numbers per Category of Invasiveness, as well as raising issues of whether ethical limits can, or should, be placed on genetic engineering. A workshop was held with the aim of bringing together Canadian animal care committee members to discuss these issues, to reflect on progress that has been made in addressing them, and to propose ways of overcoming any challenges. Although previous literature has made recommendations with regard to refinement and reduction when creating new genetically-engineered animals, the perception of the workshop participants was that some key opportunities are being missed. The participants identified the main roadblocks to the implementation of refinement and reduction alternatives as confidentiality, cost and competition. If the scientific community is to make progress concerning the implementation of refinement and reduction, particularly in the creation and use of genetically-engineered animals, addressing these roadblocks needs to be a priority.

  18. The experimental study of genetic engineering human neural stem cells mediated by lentivirus to express multigene

    Institute of Scientific and Technical Information of China (English)

    CAI Pei-qiang; TANG Xun; LIN Yue-qiu; Oudega Martin; SUN Guang-yun; XU Lin; YANG Yun-kang; ZHOU Tian-hua

    2006-01-01

    Objective:To explore the feasibility to construct genetic engineering human neural stem cells (hNSCs)mediated by lentivirus to express multigene in order to provide a graft source for further studies of spinal cord injury (SCI).Methods: Human neural stem cells from the brain cortex of human abortus were isolated and cultured, then gene was modified by lentivirus to express both green fluorescence protein (GFP) and rat neurotrophin-3(NT-3); the transgenic expression was detected by the methods of fluorescence microscope, dorsal root ganglion of fetal rats and slot blot.Results: Genetic engineering hNSCs were successfully constructed. All of the genetic engineering hNSCs which expressed bright green fluorescence were observed under the fluorescence microscope. The conditioned medium of transgenic hNSCs could induce neurite flourishing outgrowth from dorsal root ganglion (DRG). The genetic engineering hNSCs expressed high level NT-3 which could be detected by using slot blot.Conclusions: Genetic engineering hNSCs mediated by lentivirus can be constructed to express multigene successfully.

  19. Cell-matrix Interactions of Factor IX (FIX)-engineered human mesenchymal stromal cells encapsulated in RGD-alginate vs. fibrinogen-alginate microcapsules.

    Science.gov (United States)

    Sayyar, Bahareh; Dodd, Megan; Marquez-Curtis, Leah; Janowska-Wieczorek, Anna; Hortelano, Gonzalo

    2014-04-01

    The success of cell microencapsulation technology in tissue engineering and protein delivery applications depends on the viability and functionality of the encapsulated cells, which in turn are dependent upon cell/matrix interactions. In this work, we compared the viability of cord blood-derived mesenchymal stromal cells (CB MSCs), engineered to secrete factor IX (FIX) for hemophilia treatment, and encapsulated in arginine-glycine-aspartate (RGD)-alginate versus fibrinogen-alginate microcapsules. We evaluated the effect of the biomimetic matrix on cell attachment, proliferation, and secretion of FIX. Compared with nonsupplemented alginate matrix, RGD-alginate significantly enhanced the viability of the encapsulated MSCs. Further, cells in RGD-alginate displayed distinct attachment morphology, thus suggesting that RGD-alginate can potentially be used for the encapsulation of MSCs in tissue engineering applications that require enhanced cell attachment and viability. However, our data also showed that RGD-alginate microcapsules, in contrast to fibrinogen-alginate microcapsules, did not significantly improve cell proliferation of or FIX secretion by encapsulated MSCs. Our findings suggest that evidence of cell attachment alone may not accurately predict the functionality of cells in biomimetic microcapsules.

  20. Mesenchymal progenitor cells derived from synovium and infrapatellar fat pad as a source for superficial zone cartilage tissue engineering: analysis of superficial zone protein/lubricin expression.

    Science.gov (United States)

    Lee, Sang Yang; Nakagawa, Toshiyuki; Reddi, A Hari

    2010-01-01

    Superficial zone protein (SZP) is a boundary lubricant of articular cartilage in joints. As SZP at the surface of articular cartilage plays an important role in the normal function of synovial joints, the localization of SZP-secreting cells at the surface of tissue-engineered cartilage is prerequisite. The aim of this study was to identify suitable progenitor cell sources for tissue engineering of superficial zone cartilage. We investigated whether mesenchymal progenitor cells (MPCs) from synovium and infrapatellar fat pad (IFP) have the potential for secretion of SZP after chondrogenic differentiation in an aggregate pellet culture system. SZP was immunolocalized in pellets from synovium-MPCs and IFP-MPCs. The enzyme-linked immunosorbent assay analysis of SZP demonstrated that chondrogenically differentiated synovium-MPC and IFP-MPC pellets secreted SZP into media. Real-time polymerase chain reaction analysis showed significant upregulation of SZP mRNA in synovium-MPC and IFP-MPC pellets after chondrogenic differentiation. The synovium-MPCs demonstrated the higher colony-forming, proliferative, and chondrogenic potential, and exhibited greater SZP secretion after chondrogenic induction compared with IFP-MPCs. In conclusion, both synovium and IFP are promising cell sources for tissue engineering of superficial zone cartilage.

  1. An Ethical Study on the Uses of Enhancement Genetic Engineering

    Science.gov (United States)

    Kawakita, Koji

    A variety of biomedical technologies are being developed that can be used for purposes other than treating diseases. Such “enhancement technologies” can be used to improve our own and future generation's life-chances. While these technologies can help people in many ways, their use raises important ethical issues. Some arguments for anti-enhancement as well as pro-enhancement seem to rest, however, on shaky foundation. Both company engineers and the general public had better learn more from technological, economical and philosophical histories. For such subjects may provide engineers with less opportunities of technological misuses and more powers of self-esteem in addition to self-control.

  2. Genomic landscapes of endogenous retroviruses unveil intricate genetics of conventional and genetically-engineered laboratory mouse strains.

    Science.gov (United States)

    Lee, Kang-Hoon; Lim, Debora; Chiu, Sophia; Greenhalgh, David; Cho, Kiho

    2016-04-01

    Laboratory strains of mice, both conventional and genetically engineered, have been introduced as critical components of a broad range of studies investigating normal and disease biology. Currently, the genetic identity of laboratory mice is primarily confirmed by surveying polymorphisms in selected sets of "conventional" genes and/or microsatellites in the absence of a single completely sequenced mouse genome. First, we examined variations in the genomic landscapes of transposable repetitive elements, named the TREome, in conventional and genetically engineered mouse strains using murine leukemia virus-type endogenous retroviruses (MLV-ERVs) as a probe. A survey of the genomes from 56 conventional strains revealed strain-specific TREome landscapes, and certain families (e.g., C57BL) of strains were discernible with defined patterns. Interestingly, the TREome landscapes of C3H/HeJ (toll-like receptor-4 [TLR4] mutant) inbred mice were different from its control C3H/HeOuJ (TLR4 wild-type) strain. In addition, a CD14 knock-out strain had a distinct TREome landscape compared to its control/backcross C57BL/6J strain. Second, an examination of superantigen (SAg, a "TREome gene") coding sequences of mouse mammary tumor virus-type ERVs in the genomes of the 46 conventional strains revealed a high diversity, suggesting a potential role of SAgs in strain-specific immune phenotypes. The findings from this study indicate that unexplored and intricate genomic variations exist in laboratory mouse strains, both conventional and genetically engineered. The TREome-based high-resolution genetics surveillance system for laboratory mice would contribute to efficient study design with quality control and accurate data interpretation. This genetics system can be easily adapted to other species ranging from plants to humans.

  3. Genetic correction using engineered nucleases for gene therapy applications.

    Science.gov (United States)

    Li, Hongmei Lisa; Nakano, Takao; Hotta, Akitsu

    2014-01-01

    Genetic mutations in humans are associated with congenital disorders and phenotypic traits. Gene therapy holds the promise to cure such genetic disorders, although it has suffered from several technical limitations for decades. Recent progress in gene editing technology using tailor-made nucleases, such as meganucleases (MNs), zinc finger nucleases (ZFNs), TAL effector nucleases (TALENs) and, more recently, CRISPR/Cas9, has significantly broadened our ability to precisely modify target sites in the human genome. In this review, we summarize recent progress in gene correction approaches of the human genome, with a particular emphasis on the clinical applications of gene therapy.

  4. Engineering

    National Research Council Canada - National Science Library

    Includes papers in the following fields: Aerospace Engineering, Agricultural Engineering, Chemical Engineering, Civil Engineering, Electrical Engineering, Environmental Engineering, Industrial Engineering, Materials Engineering, Mechanical...

  5. Genetic engineering of novel flower colors in floricultural plants: recent advances via transgenic approaches.

    Science.gov (United States)

    Nishihara, Masahiro; Nakatsuka, Takashi

    2010-01-01

    Since the first successful genetic engineering of flower color in petunia, several new techniques have been developed and applied to modify flower color not only in model plants but also in floricultural plants. A typical example is the commercial violet-flowered carnation "Moondust series" developed by Suntry Ltd. and Florigene Ltd. More recently, blue-flowered roses have been successfully produced and are expected to be commercially available in the near future. In recent years, successful modification of flower color by sophisticated regulation of flower-pigment metabolic pathways has become possible. In this chapter, we review recent advances in flower color modification by genetic engineering, especially focusing on the methodology. We have included our own recent results on successful production of flower-color-modified transgenic plants in a model plant, tobacco and an ornamental plant, gentian. Based on these results, genetic engineering of flower color for improvement of floricultural plants is discussed.

  6. A Hybrid Neural Network-Genetic Algorithm Technique for Aircraft Engine Performance Diagnostics

    Science.gov (United States)

    Kobayashi, Takahisa; Simon, Donald L.

    2001-01-01

    In this paper, a model-based diagnostic method, which utilizes Neural Networks and Genetic Algorithms, is investigated. Neural networks are applied to estimate the engine internal health, and Genetic Algorithms are applied for sensor bias detection and estimation. This hybrid approach takes advantage of the nonlinear estimation capability provided by neural networks while improving the robustness to measurement uncertainty through the application of Genetic Algorithms. The hybrid diagnostic technique also has the ability to rank multiple potential solutions for a given set of anomalous sensor measurements in order to reduce false alarms and missed detections. The performance of the hybrid diagnostic technique is evaluated through some case studies derived from a turbofan engine simulation. The results show this approach is promising for reliable diagnostics of aircraft engines.

  7. Micropropagation, genetic engineering, and molecular biology of Populus

    Science.gov (United States)

    N. B. Klopfenstein; Y. W. Chun; M. -S. Kim; M. A. Ahuja; M. C. Dillon; R. C. Carman; L. G. Eskew

    1997-01-01

    Thirty-four Populus biotechnology chapters, written by 85 authors, are comprised in 5 sections: 1) in vitro culture (micropropagation, somatic embryogenesis, protoplasts, somaclonal variation, and germplasm preservation); 2) transformation and foreign gene expression; 3) molecular biology (molecular/genetic characterization); 4) biotic and abiotic resistance (disease,...

  8. Genetic engineering of Pichia stipitis for fermentation of xylose

    Science.gov (United States)

    Thomas W. Jeffries; N. Q. Shi; J. Y. Cho; P. Lu; K. Dahn; J. Hendrick; H. K. Sreenath

    1998-01-01

    A useful genetic system has been developed for the transformation of Pichia stipitis. This includes two selectable markers (URA3 and LEU2), integrating and autonomous replication vectors, a pop-out cassette that enables multiple targeted disruptions, and a genomic X-library for rapid cloning. Using this system we have cloned two genes for alcohol dehydrogenase (PsADH1...

  9. Potato leafroll virus : molecular analysis and genetically engineered resistance

    NARCIS (Netherlands)

    Wilk, van der F.

    1995-01-01

    The nucleotide sequence of the genomic RNA of potato leafroll virus (PLRV) was elucidated and its genetic organization deduced (Chapter 2). Six open reading frames (ORFs) were shown to be present on the genome. Both the PLRV coat protein gene and the RNA- dependent RNA polymerase gene were

  10. Non-Genetic Engineering Approaches for Isolating and Generating Novel Yeasts for Industrial Applications

    Science.gov (United States)

    Chambers, P. J.; Bellon, J. R.; Schmidt, S. A.; Varela, C.; Pretorius, I. S.

    Generating novel yeast strains for industrial applications should be quite straightforward; after all, research into the genetics, biochemistry and physiology of Baker's Yeast, Saccharomyces cerevisiae, has paved the way for many advances in the modern biological sciences. We probably know more about this humble eukaryote than any other, and it is the most tractable of organisms for manipulation using modern genetic engineering approaches. In many countries, however, there are restrictions on the use of genetically-modified organisms (GMOs), particularly in foods and beverages, and the level of consumer acceptance of GMOs is, at best, variable. Thus, many researchers working with industrial yeasts use genetic engineering techniques primarily as research tools, and strain development continues to rely on non-GM technologies. This chapter explores the non-GM tools and strategies available to such researchers.

  11. Reverse engineering gene networks: Integrating genetic perturbations with dynamical modeling

    Science.gov (United States)

    Tegnér, Jesper; Yeung, M. K. Stephen; Hasty, Jeff; Collins, James J.

    2003-01-01

    While the fundamental building blocks of biology are being tabulated by the various genome projects, microarray technology is setting the stage for the task of deducing the connectivity of large-scale gene networks. We show how the perturbation of carefully chosen genes in a microarray experiment can be used in conjunction with a reverse engineering algorithm to reveal the architecture of an underlying gene regulatory network. Our iterative scheme identifies the network topology by analyzing the steady-state changes in gene expression resulting from the systematic perturbation of a particular node in the network. We highlight the validity of our reverse engineering approach through the successful deduction of the topology of a linear in numero gene network and a recently reported model for the segmentation polarity network in Drosophila melanogaster. Our method may prove useful in identifying and validating specific drug targets and in deconvolving the effects of chemical compounds. PMID:12730377

  12. Biosensing Vibrio cholerae with Genetically Engineered Escherichia coli.

    Science.gov (United States)

    Holowko, Maciej B; Wang, Huijuan; Jayaraman, Premkumar; Poh, Chueh Loo

    2016-11-18

    Cholera is a potentially mortal, infectious disease caused by Vibrio cholerae bacterium. Current treatment methods of cholera still have limitations. Beneficial microbes that could sense and kill the V. cholerae could offer potential alternative to preventing and treating cholera. However, such V. cholerae targeting microbe is still not available. This microbe requires a sensing system to be able to detect the presence of V. cholera bacterium. To this end, we designed and created a synthetic genetic sensing system using nonpathogenic Escherichia coli as the host. To achieve the system, we have moved proteins used by V. cholerae for quorum sensing into E. coli. These sensor proteins have been further layered with a genetic inverter based on CRISPRi technology. Our design process was aided by computer models simulating in vivo behavior of the system. Our sensor shows high sensitivity to presence of V. cholerae supernatant with tight control of expression of output GFP protein.

  13. Gene therapy in dentistry: tool of genetic engineering. Revisited.

    Science.gov (United States)

    Gupta, Khushboo; Singh, Saurabh; Garg, Kavita Nitish

    2015-03-01

    Advances in biotechnology have brought gene therapy to the forefront of medical research. The concept of transferring genes to tissues for clinical applications has been discussed nearly half a century, but the ability to manipulate genetic material via recombinant DNA technology has brought this goal to reality. The feasibility of gene transfer was first demonstrated using tumour viruses. This led to development of viral and nonviral methods for the genetic modification of somatic cells. Applications of gene therapy to dental and oral problems illustrate the potential impact of this technology on dentistry. Preclinical trial results regarding the same have been very promising. In this review we will discuss methods, vectors involved, clinical implication in dentistry and scientific issues associated with gene therapy.

  14. Genetic engineering of human embryonic stem cells with lentiviral vectors.

    Science.gov (United States)

    Xiong, Chen; Tang, Dong-Qi; Xie, Chang-Qing; Zhang, Li; Xu, Ke-Feng; Thompson, Winston E; Chou, Wayne; Gibbons, Gary H; Chang, Lung-Ji; Yang, Li-Jun; Chen, Yuqing E

    2005-08-01

    Human embryonic stem (hES) cells present a valuable source of cells with a vast therapeutic potential. However, the low efficiency of directed differentiation of hES cells remains a major obstacle in their uses for regenerative medicine. While differentiation may be controlled by the genetic manipulation, effective and efficient gene transfer into hES cells has been an elusive goal. Here, we show stable and efficient genetic manipulations of hES cells using lentiviral vectors. This method resulted in the establishment of stable gene expression without loss of pluripotency in hES cells. In addition, lentiviral vectors were effective in conveying the expression of an U6 promoter-driven small interfering RNA (siRNA), which was effective in silencing its specific target. Taken together, our results suggest that lentiviral gene delivery holds great promise for hES cell research and application.

  15. Engineering and Functional Analysis of Mitotic Kinases Through Chemical Genetics.

    Science.gov (United States)

    Jones, Mathew J K; Jallepalli, Prasad V

    2016-01-01

    During mitosis, multiple protein kinases transform the cytoskeleton and chromosomes into new and highly dynamic structures that mediate the faithful transmission of genetic information and cell division. However, the large number and strong conservation of mammalian kinases in general pose significant obstacles to interrogating them with small molecules, due to the difficulty in identifying and validating those which are truly selective. To overcome this problem, a steric complementation strategy has been developed, in which a bulky "gatekeeper" residue within the active site of the kinase of interest is replaced with a smaller amino acid, such as glycine or alanine. The enlarged catalytic pocket can then be targeted in an allele-specific manner with bulky purine analogs. This strategy provides a general framework for dissecting kinase function with high selectivity, rapid kinetics, and reversibility. In this chapter we discuss the principles and techniques needed to implement this chemical genetic approach in mammalian cells.

  16. Synthetic alienation of microbial organisms by using genetic code engineering: Why and how?

    Science.gov (United States)

    Kubyshkin, Vladimir; Budisa, Nediljko

    2017-08-01

    The main goal of synthetic biology (SB) is the creation of biodiversity applicable for biotechnological needs, while xenobiology (XB) aims to expand the framework of natural chemistries with the non-natural building blocks in living cells to accomplish artificial biodiversity. Protein and proteome engineering, which overcome limitation of the canonical amino acid repertoire of 20 (+2) prescribed by the genetic code by using non-canonic amino acids (ncAAs), is one of the main focuses of XB research. Ideally, estranging the genetic code from its current form via systematic introduction of ncAAs should enable the development of bio-containment mechanisms in synthetic cells potentially endowing them with a "genetic firewall" i.e. orthogonality which prevents genetic information transfer to natural systems. Despite rapid progress over the past two decades, it is not yet possible to completely alienate an organism that would use and maintain different genetic code associations permanently. In order to engineer robust bio-contained life forms, the chemical logic behind the amino acid repertoire establishment should be considered. Starting from recent proposal of Hartman and Smith about the genetic code establishment in the RNA world, here the authors mapped possible biotechnological invasion points for engineering of bio-contained synthetic cells equipped with non-canonical functionalities. Copyright © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  17. Recent advances in the molecular genetics of resin biosynthesis and genetic engineering strategies to improve defenses in conifers

    Institute of Scientific and Technical Information of China (English)

    TANGWei

    2003-01-01

    Since the first terpenoid synthase cDNA was obtained by the reverse genetic approach from grand fir, great pro-gress in the molecular genetics of terpenoid formation has been made with angiosperms and genes encoding a monoterpene synthase, a sesquiterpene synthase, and a diterpene synthase. Tree killing bark beetles and their vectored fungal pathogens are the most destructive agents of conifer forests worldwide. Conifers defend against attack by the constitutive and inducible production of oleoresin that accumulates at the wound site to kill invaders and both flush and seal the injury. Although toxic to the bark beetle and fungal pathogen, oleoresin also plays a central role in the chemical ecology of these boring insects. Re-cent advances in the molecular genetics of terpenoid biosynthesis provide evidence for the evolutionary origins of oleoresin and permit consideration of genetic engineering strategies to improve conifer defenses as a component of modern forest bio-technology. This review described enzymes of resin biosynthesis, structural feathers of genes genomic intron and exon or-ganization, pathway organization and evolution, resin production and accumulation, interactions between conifer and bark beetle, and engineering strategies to improve conifer defenses.

  18. Simultaneous implant placement and bone regeneration around dental implants using tissue-engineered bone with fibrin glue, mesenchymal stem cells and platelet-rich plasma.

    Science.gov (United States)

    Ito, Kenji; Yamada, Yoichi; Naiki, Takahito; Ueda, Minoru

    2006-10-01

    This study was undertaken to evaluate the use of tissue-engineered bone as grafting material for alveolar augmentation with simultaneous implant placement. Twelve adult hybrid dogs were used in this study. One month after the extraction of teeth in the mandible region, bone defects on both sides of the mandible were induced using a trephine bar with a diameter of 10 mm. Dog mesenchymal stem cells (dMSCs) were obtained via iliac bone biopsy and cultured for 4 weeks before implantation. After installing the dental implants, the defects were simultaneously implanted with the following graft materials: (i) fibrin, (ii) dMSCs and fibrin (dMSCs/fibrin), (iii) dMSCs, platelet-rich plasma (PRP) and fibrin (dMSCs/PRP/fibrin) and (iv) control (defect only). The implants were assessed by histological and histomorphometric analysis, 2, 4 and 8 weeks after implantation. The implants exhibited varying degrees of bone-implant contact (BIC). The BIC was 17%, 19% and 29% (control), 20%, 22% and 25% (fibrin), 22%, 32% and 42% (dMSCs/fibrin) and 25%, 49% and 53% (dMSCs/PRP/fibrin) after 2, 4 and 8 weeks, respectively. This study suggests that tissue-engineered bone may be of sufficient quality for predictable enhancement of bone regeneration around dental implants when used simultaneous by with implant placement.

  19. Biocompatibility and enhanced osteogenic differentiation of human mesenchymal stem cells in response to surface engineered poly(D,L-lactic-co-glycolic acid) microparticles.

    Science.gov (United States)

    Rogers, Catherine M; Deehan, David J; Knuth, Callie A; Rose, Felicity R A J; Shakesheff, Kevin M; Oldershaw, Rachel A

    2014-11-01

    Tissue engineering strategies can be applied to enhancing osseous integration of soft tissue grafts during ligament reconstruction. Ligament rupture results in a hemarthrosis, an acute intra-articular bleed rich in osteogenic human mesenchymal stem cells (hMSCs). With the aim of identifying an appropriate biomaterial with which to combine hemarthrosis fluid-derived hMSCs (HF-hMSCs) for therapeutic application, this work has investigated the biocompatibility of microparticles manufactured from two forms of poly(D,L-lactic-co-glycolic acid) (PLGA), one synthesized with equal monomeric ratios of lactic acid to glycolic acid (PLGA 50:50) and the other with a higher proportion of lactic acid (PLGA 85:15) which confers a longer biodegradation time. The surfaces of both types of microparticles were functionalized by plasma polymerization with allylamine to increase hydrophilicity and promote cell attachment. HF-hMSCs attached to and spread along the surface of both forms of PLGA microparticle. The osteogenic response of HF-hMSCs was enhanced when cultured with PLGA compared with control cultures differentiated on tissue culture plastic and this was independent of the type of polymer used. We have demonstrated that surface engineered PLGA microparticles are an appropriate biomaterial for combining with HF-hMSCs and the selection of PLGA is relevant only when considering the biodegradation time for each biomedical application.

  20. On Natural Genetic Engineering: Structural Dynamism in Random Boolean Networks

    CERN Document Server

    Bull, Larry

    2012-01-01

    This short paper presents an abstract, tunable model of genomic structural change within the cell lifecycle and explores its use with simulated evolution. A well-known Boolean model of genetic regulatory networks is extended to include changes in node connectivity based upon the current cell state, e.g., via transposable elements. The underlying behaviour of the resulting dynamical networks is investigated before their evolvability is explored using a version of the NK model of fitness landscapes. Structural dynamism is found to be selected for in non-stationary environments and subsequently shown capable of providing a mechanism for evolutionary innovation when such reorganizations are inherited.

  1. Bcl-xL Genetic Modification Enhanced the Therapeutic Efficacy of Mesenchymal Stem Cell Transplantation in the Treatment of Heart Infarction

    Directory of Open Access Journals (Sweden)

    Xiaodong Xue

    2015-01-01

    Full Text Available Objectives. Low survival rate of mesenchymal stem cells (MSCs severely limited the therapeutic efficacy of cell therapy in the treatment of myocardial infarction (MI. Bcl-xL genetic modification might enhance MSC survival after transplantation. Methods. Adult rat bone marrow MSCs were modified with human Bcl-xL gene (hBcl-xL-MSCs or empty vector (vector-MSCs. MSC apoptosis and paracrine secretions were characterized using flow cytometry, TUNEL, and ELISA in vitro. In vivo, randomized adult rats with MI received myocardial injections of one of the three reagents: hBcl-xL-MSCs, vector-MSCs, or culture medium. Histochemistry, TUNEL, and echocardiography were carried out to evaluate cell engraftment, apoptosis, angiogenesis, scar formation, and cardiac functional recovery. Results. In vitro, cell apoptosis decreased 43%, and vascular endothelial growth factor (VEGF, insulin-like growth factor-1 (IGF-1, and plate-derived growth factor (PDGF increased 1.5-, 0.7-, and 1.2-fold, respectively, in hBcl-xL-MSCs versus wild type and vector-MSCs. In vivo, cell apoptosis decreased 40% and 26% in hBcl-xL-MSC group versus medium and vector-MSC group, respectively. Similar results were observed in cell engraftment, angiogenesis, scar formation, and cardiac functional recovery. Conclusions. Genetic modification of MSCs with hBcl-xL gene could be an intriguing strategy to improve the therapeutic efficacy of cell therapy in the treatment of heart infarction.

  2. Lipidomic analysis of Arabidopsis seed genetically engineered to contain DHA

    Directory of Open Access Journals (Sweden)

    Xue-Rong eZhou

    2014-09-01

    Full Text Available Metabolic engineering of omega-3 long-chain (≥C20 polyunsaturated fatty acids (ω3 LC-PUFA in oilseeds has been one of the key metabolic engineering targets in recent years. By expressing a transgenic pathway for enhancing the synthesis of the ω3 LC-PUFA docosahexaenoic acid (DHA from endogenous -linolenic acid (ALA, we obtained the production of fish oil-like proportions of DHA in Arabidopsis seed oil. Liquid chromatography-mass spectrometry (LC-MS was used to characterize the triacylglycerol (TAG, diacylglycerol (DAG and phospholipid (PL lipid classes in the transgenic and wild type Arabidopsis seeds at both developing and mature stages. The analysis identified the appearance of several abundant DHA-containing phosphatidylcholine (PC, DAG and TAG molecular species in mature seeds. The relative abundances of PL, DAG and TAG species showed a preferred combination of LC-PUFA with ALA in the transgenic seeds, where LC-PUFA were esterified in positions usually occupied by 20:1ω9. Trace amounts of di-DHA PC and tri-DHA TAG were identified, and confirmed by high resolution MS/MS. Studying the lipidome in transgenic seeds provides insights into where DHA accumulated and composed with other fatty acids of neutral and phospholipids from the developing and mature seeds.

  3. Genetic engineering in film: the case of chimeras

    Directory of Open Access Journals (Sweden)

    Josep-E. BAÑOS

    2016-04-01

    Full Text Available The development of molecular genetics in the second half of XXth century has allowed considering situations, which were in the bioscience fiction field until then. Among them, the possibility of making chimeras using the combination of genetic material is now a real option. Movies have repeatedly shown this possibility by means of literary works o directly by screen plays. This article analyzes some films that may help to understand social beliefs on chimeras in the last century. We have considered Island of lost souls (1932, The island of doctor Moreau (1977, The fly (1958, 1986, Mimic (1997 and Splice (2009. The main conclusions of this analysis are the presence of a negative view to the possibility of making chimeras following the point of view that was used in Frankenstein. The movies also lack of a consideration of the potential benefits of using chimeras. Ethical misgivings and the vision of playing God scientists avoid a impartial view of a situation, which is already among us.

  4. The hermeneutic challenge of genetic engineering: Habermas and the transhumanists.

    Science.gov (United States)

    Edgar, Andrew

    2009-06-01

    The purpose of this paper is to explore the impact that developments in transhumanist technologies may have upon human cultures (and thus upon the lifeworld), and to do so by exploring a potential debate between Habermas and the transhumanists. Transhumanists, such as Nick Bostrom, typically see the potential in genetic and other technologies for positively expanding and transcending human nature. In contrast, Habermas is a representative of those who are fearful of this technology, suggesting that it will compound the deleterious effects of the colonisation of the lifeworld, further constraining human autonomy and undermining the meaningfulness of the lifeworld by expanding the technological control and manipulation of humanity. It will be argued that these opposed positions are grounded in fundamentally different understandings of the consequences of scientific and technological advance. On one level, the transhumanists remain confident that the lifeworld has within it the resources necessary to find meaning and purpose in a society deeply infused by genetic technology. Habermas disagrees. On another level, the difference is articulated by Horkheimer and Adorno in Dialectic of Enlightenment, primarily by challenging what may be understood as a Baconian faith in science as a project for the domination of nature (where nature is an infinitely malleable material, to be dominated and shaped, without adverse consequences, purely for the purposes of human survival). While the transhumanists broadly embrace this faith, Habermas returns to something akin to Horkheimer and Adorno's pessimistic scepticism.

  5. Projecting potential adoption of genetically engineered freeze-tolerant Eucalyptus in the United States

    Science.gov (United States)

    David N. Wear; Ernest Dixon IV; Robert C. Abt; Navinder Singh

    2015-01-01

    Development of commercial Eucalyptus plantations has been limited in the United States because of the species’ sensitivity to freezing temperatures. Recently developed genetically engineered clones of a Eucalyptus hybrid, which confer freeze tolerance, could expand the range of commercial plantations. This study explores how...

  6. Rapid engineering of versatile molecular logic gates using heterologous genetic transcriptional modules.

    Science.gov (United States)

    Wang, Baojun; Buck, Martin

    2014-10-11

    We designed and constructed versatile modular genetic logic gates in bacterial cells. These function as digital logic 1-input Buffer gate, 2-input and 3-input AND gates with one inverted input and integrate multiple chemical input signals in customised logic manners. Such rapidly engineered devices serve to achieve increased sensing signal selectivity.

  7. 77 FR 41350 - Monsanto Co.; Determination of Nonregulated Status of Soybean Genetically Engineered To Produce...

    Science.gov (United States)

    2012-07-13

    ... article under our regulations governing the introduction of certain genetically engineered organisms. Our....aphis.usda.gov/biotechnology/not_reg.html and are posted with the previous notice and the comments we..., Biotechnology Regulatory Services, APHIS, 4700 River Road Unit 147, Riverdale, MD 20737-1236; (301) 851-3954...

  8. 76 FR 78232 - Monsanto Co.; Determination of Nonregulated Status for Soybean Genetically Engineered To Have a...

    Science.gov (United States)

    2011-12-16

    ... the introduction of certain genetically engineered organisms. Our determination is based on our.../biotechnology/not_reg.html and are posted with the previous notice and the comments we received on the... INFORMATION CONTACT: Mr. Evan Chestnut, Policy Analyst, Biotechnology Regulatory Services, APHIS, 4700 River...

  9. Genetic engineering of plant volatile terpenoids: effects on a herbivore, a predator and a parasitoid

    NARCIS (Netherlands)

    Kos, M.; Houshyani, B.; Overeem, A.J.; Bouwmeester, H.J.; Weldegergis, B.T.; van Loon, J.J.A.; Dicke, M.; Vet, L.E.M.

    2013-01-01

    BACKGROUND: Most insect-resistant transgenic crops employ toxins to control pests. A novel approach is to enhance the effectiveness of natural enemies by genetic engineering of the biosynthesis of volatile organic compounds (VOCs). Before the commercialisation of such transgenic plants can be pursue

  10. History and future of genetically engineered food animal regulation: an open request.

    Science.gov (United States)

    Wells, Kevin D

    2016-06-01

    Modern biotechnology resulted from of a series of incremental improvements in the understanding of DNA and the enzymes that nature evolved to manipulate it. As the potential impact of genetic engineering became apparent, scientists began the process of trying to identify the potential unintended consequences. Restrictions to recombinant DNA experimentation were at first self-imposed. Collaborative efforts between scientists and lawyers formalized an initial set of guidelines. These guidelines have been used to promulgate regulations around world. However, the initial guidelines were only intended as a starting point and were motivated by a specific set of concerns. As new data became available, the guidelines and regulations should have been adapted to the new knowledge. Instead, other social drivers drove the development of regulations. For most species and most applications, the framework that was established has slowly allowed some products to reach the market. However, genetically engineered livestock that are intended for food have been left in a regulatory state of limbo. To date, no genetically engineered food animal is available in the marketplace. A short history and a U.S.-based genetic engineer's perspective are presented. In addition, a request to regulatory agencies is presented for consideration as regulation continues to evolve. Regulators appear to have shown preference for the slow, random progression of evolution over the efficiency of intentional design.

  11. 'HoneySweet' plum - a valuable genetically engineered fruit-tree cultivar and germplasm resource

    Science.gov (United States)

    ‘HoneySweet’ is a plum variety developed through genetic engineering to be highly resistant to plum pox potyvirus (PPV), the causal agent of sharka disease, that threatens stone-fruit industries world-wide and most specifically, in Europe. Field testing for over 15 years in Europe has demonstrated ...

  12. Development of enzymes and enzyme systems by genetic engineering to convert biomass to sugars

    Science.gov (United States)

    TITLE Development of Enzymes and Enzyme Systems by Genetic Engineering to Convert Biomass to Sugars ABSTRACT Plant cellulosic material is one of the most viable renewable resources for the world’s fuel and chemical feedstock needs. Currently ethanol derived from corn starch is the most common li...

  13. Enhancing the Internationalisation of Distance Education in the Biological Sciences: The DUNE Project and Genetic Engineering.

    Science.gov (United States)

    Leach, C. K.; And Others

    1997-01-01

    Describes the Distance Educational Network of Europe (DUNE) project that aims at enhancing the development of distance education in an international context. Highlights issues relating to the delivery of distance-learning courses in a transnational forum. Describes the genetic engineering course that aims at explaining the core techniques of…

  14. Between creation, evolution and genetic engineering: biology in need of a new bioethics?

    NARCIS (Netherlands)

    Gupta, J.A.

    2009-01-01

    Technological interventions into biological processes through genetic engineering in the twenty-fi rst century could speed up evolution at the velocity of light years in comparison with the millions of years it took for Homo sapiens to reach this stage of evolution until this new millennium. Will th

  15. Genetically engineered alfalfa and feral alfalfa plants: What should growers know?

    Science.gov (United States)

    Alfalfa (Medicago sativa subsp. sativa L) is the world’s most important forage crop. The western United States is the most important production area for both alfalfa forage and alfalfa seed. Alfalfa was the first major perennial genetically-engineered (GE)crop and a GE trait for resistance to glypho...

  16. Conversion of Glycerol to 3-Hydroxypropanoic Acid by Genetically Engineered Bacillus subtilis

    DEFF Research Database (Denmark)

    Kalantari, Aida; Chen, Tao; Ji, Boyang

    2017-01-01

    of glycerol into 3-HP. Our recombinant B. subtilis strains overexpress the two-step heterologous pathway containing glycerol dehydratase and aldehyde dehydrogenase from K. pneumoniae. Genetic engineering, driven by in silico optimization, and optimization of cultivation conditions resulted in a 3-HP titer...

  17. A field release of genetically engineered gypsy moth (Lymantria dispar L.) Nuclear Polyhedrosis Virus (LdNPV)

    Science.gov (United States)

    Vincent D' Amico; Joseph S. Elkinton; John D. Podgwaite; James M. Slavicek; Michael L. McManus; John P. Burand

    1999-01-01

    The gypsy moth (Lymantria dispar L.) nuclear polyhedrosis virus was genetically engineered for nonpersistence by removal of the gene coding for polyhedrin production and stabilized using a coocclusion process. A β-galactosidase marker gene was inserted into the genetically engineered virus (LdGEV) so that infected larvae could be tested for...

  18. [Progress of research on genetic engineering antibody and its application in prevention and control of parasitic diseases].

    Science.gov (United States)

    Yao, Yuan; Yu, Chuan-xin

    2013-08-01

    Antibody has extensive application prospects in the biomedical field. The inherent disadvantages of traditional polyclonal antibody and monoclonal antibody limit their application values. The humanized and fragmented antibody remodeling has given a rise to a series of genetic engineered antibody variant. This paper reviews the progress of research on genetic engineering antibody and its application in prevention and control of parasitic diseases.

  19. Genetically engineered biological agents in therapy for systemic lupus erythematosus

    Directory of Open Access Journals (Sweden)

    Elena Aleksandrovna Aseeva

    2013-01-01

    Full Text Available Systemic lupus erythematosus (SLE is a prototype for chronic autoimmune disease. Its prevalence is 20 to 70 cases per 100,000 women and varies by race and ethnicity. Despite considerable progress in traditional therapy, many problems associated with the management of these patients need to be immediately solved: thus, 50-80% are found to have activity signs and/or frequent exacerbations and about 30% of the patients have to stop work; Class IV lupus nephritis increases the risk of terminalrenal failure. In the past 20 years great progress has been made in studying the pathogenesis of SLE: biological targets to affect drugs have been sought and fundamentally new therapeutic goals defined. Belimumab is the first genetically biological agent specially designed to treat SLE, which is rightly regarded as one of the most important achievements of rheumatology in the past 50 years.

  20. Human Wharton's jelly-derived mesenchymal stromal cells engineered to secrete Epstein-Barr virus interleukin-10 show enhanced immunosuppressive properties.

    Science.gov (United States)

    Quaranta, Paola; Focosi, Daniele; Di Iesu, Marilena; Cursi, Chiara; Zucca, Alessandra; Curcio, Michele; Lapi, Simone; Boldrini, Linda; Stampacchia, Giulia; Paolicchi, Aldo; Scatena, Fabrizio; Freer, Giulia; Pistello, Mauro

    2016-02-01

    Mesenchymal stromal cells (MSCs) modulate the immune response and represent a potential treatment for inflammatory and autoimmune diseases. We hypothesized that this feature could be potentiated by co-administering anti-inflammatory cytokines. In this article, we asked whether engineering of Wharton Jelly-derived human MSCs (WJ-hMSCs) to express an anti-inflammatory cytokine increases cell immunomodulatory properties without altering their native features. We used Epstein-Barr virus-derived interleukin-10 (vIL-10), which shares some immunosuppressive properties with human IL-10 but lacks immunostimulatory activity. Engineering was accomplished by transducing WJ-hMSCs with a self-inactivating feline immunodeficiency virus-derived vector co-expressing vIL-10 and herpes simplex virus type-1 thymidine kinase (TK). TK was added to allow future tracking of WJ-hMSC in vivo by positron electron tomography (PET). The results show that (i) expression of TK and/or vIL-10 does not change WJ-hMSC phenotypic and functional properties; (ii) vIL-10 is secreted, biologically active and enhances the immunosuppressing functions of WJ-hMSCs; (iii) v-IL10 and TK can be produced simultaneously by the same cells and do not interfere with each other. WJ-hMSCs engineered to secrete vIL-10 could be a powerful tool for adoptive cell therapy of immune-mediated diseases, and therefore, additional studies are warranted to confirm their efficacy in suitable animal disease models. Copyright © 2015 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

  1. The effect of genetically engineered glucagon on glucose recovery after hypoglycaemia in man

    DEFF Research Database (Denmark)

    Hvidberg, A; Jørgensen, S; Hilsted, J

    1992-01-01

    To compare the effect on glucose recovery after insulin-induced hypoglycaemia of intramuscular genetically engineered glucagon, intramuscular glucagon from pancreatic extraction and intravenous glucose, we examined 10 healthy subjects during blockage of glucose counterregulation with somatostatin...... appearance rate were far more protracted after i.m. glucagon than after i.v. glucose. These results suggest that genetically engineered glucagon and glucagon from pancreatic extraction have a similar effect on hepatic glucose production rate. Due to the protracted effect of intramuscular glucagon, a combined......, propranolol and phentolamine. Each subject was studied on three separate occasions. Thirty min after a bolus injection of 0.075 iu soluble insulin per kilogram body weight the subjects received one of the following treatments: 1 mg glucagon from pancreatic extraction intramuscularly; 1 mg genetically...

  2. A Candida albicans CRISPR system permits genetic engineering of essential genes and gene families.

    Science.gov (United States)

    Vyas, Valmik K; Barrasa, M Inmaculada; Fink, Gerald R

    Candida albicans is a pathogenic yeast that causes mucosal and systematic infections with high mortality. The absence of facile molecular genetics has been a major impediment to analysis of pathogenesis. The lack of meiosis coupled with the absence of plasmids makes genetic engineering cumbersome, especially for essential functions and gene families. We describe a C. albicans CRISPR system that overcomes many of the obstacles to genetic engineering in this organism. The high frequency with which CRISPR-induced mutations can be directed to target genes enables easy isolation of homozygous gene knockouts, even without selection. Moreover, the system permits the creation of strains with mutations in multiple genes, gene families, and genes that encode essential functions. This CRISPR system is also effective in a fresh clinical isolate of undetermined ploidy. Our method transforms the ability to manipulate the genome of Candida and provides a new window into the biology of this pathogen.

  3. Cell sourcing for bone tissue engineering: amniotic fluid stem cells have a delayed, robust differentiation compared to mesenchymal stem cells.

    Science.gov (United States)

    Peister, Alexandra; Woodruff, Maria A; Prince, Jarod J; Gray, Derwin P; Hutmacher, Dietmar W; Guldberg, Robert E

    2011-07-01

    Cell based therapies for bone regeneration are an exciting emerging technology, but the availability of osteogenic cells is limited and an ideal cell source has not been identified. Amniotic fluid-derived stem cells (AFS) and bone-marrow derived mesenchymal stem cells (MSCs) were compared to determine their osteogenic differentiation capacity in both 2D and 3D environments. In 2D culture, the AFS cells produced more mineralized matrix but delayed peaks in osteogenic markers. Cells were also cultured on 3D scaffolds constructed of poly-ε-caprolactone for 15 weeks. MSCs differentiated more quickly than AFS cells on 3D scaffolds, but mineralized matrix production slowed considerably after 5 weeks. In contrast, the rate of AFS cell mineralization continued to increase out to 15 weeks, at which time AFS constructs contained 5-fold more mineralized matrix than MSC constructs. Therefore, cell source should be taken into consideration when used for cell therapy, as the MSCs would be a good choice for immediate matrix production, but the AFS cells would continue robust mineralization for an extended period of time. This study demonstrates that stem cell source can dramatically influence the magnitude and rate of osteogenic differentiation in vitro. Copyright © 2011 Elsevier B.V. All rights reserved.

  4. Genetic engineering in agriculture and corporate engineering in public debate: risk, public relations, and public debate over genetically modified crops.

    Science.gov (United States)

    Patel, Rajeev; Torres, Robert J; Rosset, Peter

    2005-01-01

    Corporations have long influenced environmental and occupational health in agriculture, doing a great deal of damage, making substantial profits, and shaping public debate to make it appear that environmental misfortunes are accidents of an otherwise well-functioning system, rather than systemic. The debate over the genetically modified (GM) crops is an example. The largest producer of commercial GM seeds, Monsanto, exemplifies the industry's strategies: the invocation of poor people as beneficiaries, characterization of opposition as technophobic or anti-progress, and portrayal of their products as environmentally beneficial in the absence of or despite the evidence. This strategy is endemic to contemporary market capitalism, with its incentives to companies to externalize health and environmental costs to increase profits.

  5. Antinociceptive Effect of Intrathecal Injection of Genetically Engineered Human Bone Marrow Stem Cells Expressing the Human Proenkephalin Gene in a Rat Model of Bone Cancer Pain

    Directory of Open Access Journals (Sweden)

    Yi Sun

    2017-01-01

    Full Text Available Background. This study aimed to investigate the use of human bone marrow mesenchymal stem cells (hBMSCs genetically engineered with the human proenkephalin (hPPE gene to treat bone cancer pain (BCP in a rat model. Methods. Primary cultured hBMSCs were passaged and modified with hPPE, and the cell suspensions (6 × 106 were then intrathecally injected into a rat model of BCP. Paw mechanical withdrawal threshold (PMWT was measured before and after BCP. The effects of hPPE gene transfer on hBMSC bioactivity were analyzed in vitro and in vivo. Results. No changes were observed in the surface phenotypes and differentiation of hBMSCs after gene transfer. The hPPE-hBMSC group showed improved PMWT values on the ipsilateral side of rats with BCP from day 12 postoperatively, and the analgesic effect was reversed by naloxone. The levels of proinflammatory cytokines such as IL-1β and IL-6 were ameliorated, and leucine-enkephalin (L-EK secretion was augmented, in the hPPE-engineered hBMSC group. Conclusion. The intrathecal administration of BMSCs modified with the hPPE gene can effectively relieve pain caused by bone cancer in rats and might be a potentially therapeutic tool for cancer-related pain in humans.

  6. Antinociceptive Effect of Intrathecal Injection of Genetically Engineered Human Bone Marrow Stem Cells Expressing the Human Proenkephalin Gene in a Rat Model of Bone Cancer Pain

    Science.gov (United States)

    Tian, Yuke; Li, Haifeng; Zhang, Dengwen; Sun, Qiang

    2017-01-01

    Background. This study aimed to investigate the use of human bone marrow mesenchymal stem cells (hBMSCs) genetically engineered with the human proenkephalin (hPPE) gene to treat bone cancer pain (BCP) in a rat model. Methods. Primary cultured hBMSCs were passaged and modified with hPPE, and the cell suspensions (6 × 106) were then intrathecally injected into a rat model of BCP. Paw mechanical withdrawal threshold (PMWT) was measured before and after BCP. The effects of hPPE gene transfer on hBMSC bioactivity were analyzed in vitro and in vivo. Results. No changes were observed in the surface phenotypes and differentiation of hBMSCs after gene transfer. The hPPE-hBMSC group showed improved PMWT values on the ipsilateral side of rats with BCP from day 12 postoperatively, and the analgesic effect was reversed by naloxone. The levels of proinflammatory cytokines such as IL-1β and IL-6 were ameliorated, and leucine-enkephalin (L-EK) secretion was augmented, in the hPPE-engineered hBMSC group. Conclusion. The intrathecal administration of BMSCs modified with the hPPE gene can effectively relieve pain caused by bone cancer in rats and might be a potentially therapeutic tool for cancer-related pain in humans. PMID:28286408

  7. Genetically engineered multivalent single chain antibody constructs for cancer therapy

    Energy Technology Data Exchange (ETDEWEB)

    Surinder Batra, Ph D

    2006-02-27

    its tumor: normal tissue ratio for improved therapeutic index, we engineered a variety antibody constructs. These constructs were evaluated using novel approaches like special radionuclides, pretargeting and optimization. Due to the smaller size, the engineered antibody molecules should penetrate better throughout a tumor mass, with less dose heterogeneity, than is the case with intact IgG. Multivalent scFvs with an appropriate radionuclide, therefore, hold promising prospects for cancer therapy and clinical imaging in MAb-based radiopharmaceuticals. In addition, the human anti-mouse antibodies (HAMA) responses in patients against antibody-based therapy are usually directed against the immunoglobulin constant regions; however, anti-idiotypic responses can also be detected. The HAMA responses reduce the efficacy of treatment by removing the circulating antibody molecules, fragments, and possibly scFvs by altering the pharmacokinetic properties of the antibody. HAMA responses against divalent IgG, divalent Ig fragments, and possibly multimeric scFvs could cause immune complex formation with hypersensitivity or allergic reactions that could be harmful to patients. The use of small molecules, such as scFvs (monomeric as well as multimeric), with their shorter biological half-lives and the lack of the constant regions and humanized variable (binding regions) performed in our studies should reduce the development of HAMA. The generation of humanized and fully human scFvs should further reduce the development of HAMA. Specific accomplishments on the project are the production of large amounts of recombinant antibodies as they are required in large amounts for cancer diagnosis and therapy. A variety of single-chain Fv (scFv) constructs were engineered for the desired pharmacokinetic properties. Tetrameric and dimeric scFvs showed a two-fold advantage: (1) there was a considerable gain in avidity as compared to smaller fragments, and (2) the biological half-life was more

  8. Osteoblastic Differentiation of Human and Equine Adult Bone Marrow-Derived Mesenchymal Stem Cells when BMP-2 or BMP-7 homodimer genetic modification is compared to BMP-2/7 heterodimer genetic modification in the Presence and Absence of Dexamethasone

    Science.gov (United States)

    Carpenter, RS; Goodrich, LR; Frisbie, DD; Kisiday, JD; Carbone, B; McIlwraith, CW; Centeno, CJ; Hidaka, C

    2010-01-01

    Bone marrow-derived mesenchymal stem cells (BMDMSCs) have been targeted for use in enhancement of bone healing; and their osteogenic potential may be further augmented by genes encoding bone morphogenetic proteins (BMP’s). The purpose of this study was to compare the effect of genetic modification of human and equine BMDMSCs with BMP-2 or 7 or BMP-2 and 7 on their osteoblastogenic differentiation in the presence or absence of dexamethasone. The BMDMSCs were harvested from the iliac crest of 3 human donors and tuber coxae of 3 equine donors. Monolayer cells were genetically modified using adenovirus vectors encoding BMP-2, -7 or both and cultured in the presence or absence of dexamethasone. Expression of BMPs was confirmed by enzyme linked immunosorbent assay. To evaluate osteoblastic differentiation, cellular morphology was assessed every other day and expression and secretion of alkaline phosphatase (ALP), as well as expression levels of osteonectin, osteocalcin, and Runx2 were measured for up to 14 days. Human and equine BMDMSCs showed a capacity for osteogenic differentiation regardless of genetic modification or dexamethasone supplementation. Dexamethasone supplementation was more important for osteoblastogenic differentiation of equine BMDMSCs than human BMDMSCs. Genetic modification of BMDMSCs increased ALP secretion with AdBMP-2 homodimer having the greatest effect in both human and equine cells compared to AdBMP 7 or AdBMP 2/7. BMP protein elution rates reached their maximal concentration between day 4 and 8 and remained relatively stable thereafter, suggesting that genetically modified BMDMSCs could be useful for cell-based delivery of BMPs to a site of bone formation. PMID:20309952

  9. From Precaution to Peril: Public Relations Across Forty Years of Genetic Engineering.

    Science.gov (United States)

    Hogan, Andrew J

    2016-12-01

    The Asilomar conference on genetic engineering in 1975 has long been pointed to by scientists as a model for internal regulation and public engagement. In 2015, the organizers of the International Summit on Human Gene Editing in Washington, DC looked to Asilomar as they sought to address the implications of the new CRISPR gene editing technique. Like at Asilomar, the conveners chose to limit the discussion to a narrow set of potential CRISPR applications, involving inheritable human genome editing. The adoption by scientists in 2015 of an Asilomar-like script for discussing genetic engineering offers historians the opportunity to analyze the adjustments that have been made since 1975, and to identify the blind spots that remain in public engagement. Scientists did take important lessons from the fallout of their limited engagement with public concerns at Asilomar. Nonetheless, the scientific community has continued to overlook some of the longstanding public concerns about genetic engineering, in particular the broad and often covert genetic modification of food products. Copyright © 2016 Elsevier Ltd. All rights reserved.

  10. Mesenchymal stem cells: Identification, phenotypic characterization, biological properties and potential for regenerative medicine through biomaterial micro-engineering of their niche.

    Science.gov (United States)

    Kobolak, Julianna; Dinnyes, Andras; Memic, Adnan; Khademhosseini, Ali; Mobasheri, Ali

    2016-04-15

    Mesenchymal stem cells (MSCs) are multipotent stem cells. Although they were originally identified in bone marrow and described as 'marrow stromal cells', they have since been identified in many other anatomical locations in the body. MSCs can be isolated from bone marrow, adipose tissue, umbilical cord and other tissues but the richest tissue source of MSCs is fat. Since they are adherent to plastic, they may be expanded in vitro. MSCs have a distinct morphology and express a specific set of CD (cluster of differentiation) molecules. The phenotypic pattern for the identification of MSCs cells requires expression of CD73, CD90, and CD105 and lack of CD34, CD45, and HLA-DR antigens. Under appropriate micro-environmental conditions MSCs can proliferate and give rise to other cell types. Therefore, they are ideally suited for the treatment of systemic inflammatory and autoimmune conditions. They have also been implicated as key players in regenerating injured tissue following injury and trauma. MSC populations isolated from adipose tissue may also contain regulatory T (Treg) cells, which have the capacity for modulating the immune system. The immunoregulatory and regenerative properties of MSCs make them ideal for use as therapeutic agents in vivo. In this paper we review the literature on the identification, phenotypic characterization and biological properties of MSCs and discuss their potential for applications in cell therapy and regenerative medicine. We also discuss strategies for biomaterial micro-engineering of the stem cell niche.

  11. Application potential of bone marrow mesenchymal stem cell (BMSCs) based tissue-engineering for spinal cord defect repair in rat fetuses with spina bifida aperta.

    Science.gov (United States)

    Li, Xiaoshuai; Yuan, Zhengwei; Wei, Xiaowei; Li, Hui; Zhao, Guifeng; Miao, Jiaoning; Wu, Di; Liu, Bo; Cao, Songying; An, Dong; Ma, Wei; Zhang, Henan; Wang, Weilin; Wang, Qiushi; Gu, Hui

    2016-04-01

    Spina bifida aperta are complex congenital malformations resulting from failure of fusion in the spinal neural tube during embryogenesis. Despite surgical repair of the defect, most patients who survive with spina bifida aperta have a multiple system handicap due to neuron deficiency of the defective spinal cord. Tissue engineering has emerged as a novel treatment for replacement of lost tissue. This study evaluated the prenatal surgical approach of transplanting a chitosan-gelatin scaffold seeded with bone marrow mesenchymal stem cells (BMSCs) in the healing the defective spinal cord of rat fetuses with retinoic acid induced spina bifida aperta. Scaffold characterisation revealed the porous structure, organic and amorphous content. This biomaterial promoted the adhesion, spreading and in vitro viability of the BMSCs. After transplantation of the scaffold combined with BMSCs, the defective region of spinal cord in rat fetuses with spina bifida aperta at E20 decreased obviously under stereomicroscopy, and the skin defect almost closed in many fetuses. The transplanted BMSCs in chitosan-gelatin scaffold survived, grew and expressed markers of neural stem cells and neurons in the defective spinal cord. In addition, the biomaterial presented high biocompatibility and slow biodegradation in vivo. In conclusion, prenatal transplantation of the scaffold combined with BMSCs could treat spinal cord defect in fetuses with spina bifida aperta by the regeneration of neurons and repairmen of defective region.

  12. Novel biomimetic tripolymer scaffolds consisting of chitosan, collagen type 1, and hyaluronic acid for bone marrow-derived human mesenchymal stem cells-based bone tissue engineering.

    Science.gov (United States)

    Mathews, Smitha; Bhonde, Ramesh; Gupta, Pawan Kumar; Totey, Satish

    2014-11-01

    Human bone marrow-derived mesenchymal stem cells (hMSCs) are an ideal osteogenic cell source for bone tissue engineering (BTE). A scaffold, in the context of BTE, is the extracellular matrix (ECM) that provides the unique microenvironment and play significant role in regulating cell behavior, differentiation, and development in an in vitro culture system. In this study, we have developed novel biomimetic tripolymer scaffolds for BTE using an ECM protein, collagen type 1; an ECM glycosaminoglycan, hyaluronic acid; and a natural osteoconductive polymer, chitosan. The scaffolds were characterized by scanning electron microscopy (SEM) and swelling ratio. The scaffolds were seeded with hMSCs and tested for cytocompatibility and osteogenic potential. The scaffolds supported cell adhesion, enhanced cell proliferation, promoted cell migration, showed good cell viability, and osteogenic potential. The cells were able to migrate out from the scaffolds in favorable conditions. SEM, alkaline phosphatase assay, and immunofluorescent staining confirmed the differentiation of hMSCs to osteogenic lineage in the scaffolds. In conclusion, we have successfully developed biomimetic scaffolds that supported the proliferation and differentiation of hMSCs. These scaffolds hold great promise as a cell-delivery vehicle for regenerative therapies and as a support system for enhancing bone regeneration. © 2014 Wiley Periodicals, Inc.

  13. Ethical Analysis of Genetic Engineering%基因工程的伦理分析

    Institute of Scientific and Technical Information of China (English)

    周青龙

    2013-01-01

    Since 1977, American scientists in the world since the first time genetically engineered to produce human growth hormone, genetic engineering continues to bear the fruit of fruitful gratifying. Now, genetic engineering has been widely applied in all aspects of society. However, the science and technology is a double-edged sword. With the rapid development of genetic engineering technology, it is also produced many negative effects, so that people have to produce all kinds of worries and anxieties. Genetic engineering without restriction left unchecked, will occur contrary to the laws of nature and ethical issues, it will bring disaster to human society, resulting in consequences? Genetic engineering services for the peace and progress of mankind, must step up to the norms, moral constraints, thereby establishing public international law, so that the great discovery and shocking change comes to the change of the legal system.%  自1977年美国科学家在世界上首次用基因工程生产人类生长激素以来,基因工程不断结出令人欣喜的丰硕之果。现如今,基因工程已广泛应用在社会的各个方面。然而,科学技术是把双刃剑。随着基因工程这一技术的迅猛发展,其也产生了许多负面影响,使人们不得不产生各种担心和忧虑。基因工程如不加限制地任其发展,会不会发生违背自然规律和伦理道德的问题,会不会给人类社会带来灾难,造成恶果?基因工程要为人类和平与进步服务,就必须加紧以规范,进行道德约束,进而建立国际公法,以便让伟大的发现、震撼的变革,走向更改化、法制化。

  14. Construction of engineering adipose-like tissue in vivo utilizing human insulin gene-modified umbilical cord mesenchymal stromal cells with silk fibroin 3D scaffolds.

    Science.gov (United States)

    Li, Shi-Long; Liu, Yi; Hui, Ling

    2015-12-01

    We evaluated the use of a combination of human insulin gene-modified umbilical cord mesenchymal stromal cells (hUMSCs) with silk fibroin 3D scaffolds for adipose tissue engineering. In this study hUMSCs were isolated and cultured. HUMSCs infected with Ade-insulin-EGFP were seeded in fibroin 3D scaffolds with uniform 50-60 µm pore size. Silk fibroin scaffolds with untransfected hUMSCs were used as control. They were cultured for 4 days in adipogenic medium and transplanted under the dorsal skins of female Wistar rats after the hUMSCs had been labelled with chloromethylbenzamido-1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate (CM-Dil). Macroscopical impression, fluorescence observation, histology and SEM were used for assessment after transplantation at 8 and 12 weeks. Macroscopically, newly formed adipose tissue was observed in the experimental group and control group after 8 and 12 weeks. Fluorescence observation supported that the formed adipose tissue originated from seeded hUMSCs rather than from possible infiltrating perivascular tissue. Oil red O staining of newly formed tissue showed that there was substantially more tissue regeneration in the experimental group than in the control group. SEM showed that experimental group cells had more fat-like cells, whose volume was larger than that of the control group, and degradation of the silk fibroin scaffold was greater under SEM observation. This study provides significant evidence that hUMSCs transfected by adenovirus vector have good compatibility with silk fibroin scaffold, and adenoviral transfection of the human insulin gene can be used for the construction of tissue-engineered adipose. Copyright © 2013 John Wiley & Sons, Ltd.

  15. Evaluation of bone marrow derived mesenchymal stem cells for full-thickness wound healing in comparison to tissue engineered chitosan scaffold in rabbit.

    Science.gov (United States)

    Rajabian, Mohammad Hossein; Ghorabi, Gholam Hossein; Geramizadeh, Bita; Sameni, Safoura; Ayatollahi, Maryam

    2017-02-01

    Chronic wounds present a major challenge in modern medicine. Even under optimal conditions, the healing process may lead to scarring and fibrosis. The ability of mesenchymal stem cells (MSCs) to differentiate into other cell types makes these cells an attractive therapeutic tool for cell transplantation. Both tissue-engineered construct and MSC therapy are among the current wound healing procedures and potential care. Chitosan has been widely applied in tissue engineering because of its biocompatibility and biodegradability. The aim of the current work was to compare the efficiency of MSCs and chitosan dressing, alone or in combination treatment on wound healing. This study was conducted on 15 rabbits, which were randomly divided in 3 groups based on the type of treatment with MSCs, chitosan dressing and combination of both. A full-thickness skin defect was excised from the right and left side of the back of each animals. Defects on right sides were filled with treatments and left side defects were left as control. Evaluation of the therapeutic effectiveness was performed through a variety of clinical and microscopical evaluations and measurements of the process of wound healing on days 7, 14, 21, and 28. Histological evaluation of wound healing was classified by different scoring systems. The data indicated that wounds treated with bone marrow derived MSC had enhanced cellularity and better epidermal regeneration. During the early stages of wound healing, the closure rate of bone marrow derived MSC-treated wounds were significantly higher than other treatments (Pchitosan treatment was slower than the control group. This study revealed advanced granulation tissue formation and epithelialization in wounds treated with MSCs, and may suggests this treatment as an effective applicant in wound healing process. Chitosan scaffold dressings, whether alone or in combination with MSCs, have worsened the wound healing as compared to the control group. Copyright © 2016

  16. Umbilical cord Wharton's jelly repeated culture system: a new device and method for obtaining abundant mesenchymal stem cells for bone tissue engineering.

    Directory of Open Access Journals (Sweden)

    Zhengqi Chang

    Full Text Available To date, various types of cells for seeding regenerative scaffolds have been used for bone tissue engineering. Among seed cells, the mesenchymal stem cells derived from human umbilical cord Wharton's jelly (hUCMSCs represent a promising candidate and hold potential for bone tissue engineering due to the the lack of ethical controversies, accessibility, sourced by non-invasive procedures for donors, a reduced risk of contamination, osteogenic differentiation capacities, and higher immunomodulatory capacity. However, the current culture methods are somewhat complicated and inefficient and often fail to make the best use of the umbilical cord (UC tissues. Moreover, these culture processes cannot be performed on a large scale and under strict quality control. As a result, only a small quantity of cells can be harvested using the current culture methods. To solve these problems, we designed and evaluated an UC Wharton's jelly repeated culture device. Using this device, hUCMSCs were obtained from the repeated cultures and their quantities and biological characteristics were compared. We found that using our culture device, which retained all tissue blocks on the bottom of the dish, the total number of obtained cells increased 15-20 times, and the time required for the primary passage was reduced. Moreover, cells harvested from the repeated cultures exhibited no significant difference in their immunophenotype, potential for multilineage differentiation, or proliferative, osteoinductive capacities, and final osteogenesis. The application of the repeated culture frame (RCF not only made full use of the Wharton's jelly but also simplified and specified the culture process, and thus, the culture efficiency was significantly improved. In summary, abundant hUCMSCs of dependable quality can be acquired using the RCF.

  17. Mesenchymal stem cells as therapeutic delivery vehicles targeting tumor stroma

    DEFF Research Database (Denmark)

    Serakinci, Nedime; Christensen, Rikke; Sørensen, Flemming Brandt

    2011-01-01

    The field of stem cell biology continues to evolve by characterization of further types of stem cells and by exploring their therapeutic potential for experimental and clinical applications. Human mesenchymal stem cells (hMSCs) are one of the most promising candidates simply because...... better understanding and in vivo supporting data. The homing ability of hMSCs was investigated by creating a human xenograft model by transplanting an ovarian cancer cell line into immunocompromised mice. Then, genetically engineered hMSC-telo1 cells were injected through the tail vein...

  18. Role of stem cells in large animal genetic engineering in the TALENs-CRISPR era.

    Science.gov (United States)

    Park, Ki-Eun; Telugu, Bhanu Prakash V L

    2013-01-01

    The establishment of embryonic stem cells (ESCs) and gene targeting technologies in mice has revolutionised the field of genetics. The relative ease with which genes can be knocked out, and exogenous sequences introduced, has allowed the mouse to become the prime model for deciphering the genetic code. Not surprisingly, the lack of authentic ESCs has hampered the livestock genetics field and has forced animal scientists into adapting alternative technologies for genetic engineering. The recent discovery of the creation of induced pluripotent stem cells (iPSCs) by upregulation of a handful of reprogramming genes has offered renewed enthusiasm to animal geneticists. However, much like ESCs, establishing authentic iPSCs from the domestic animals is still beset with problems, including (but not limited to) the persistent expression of reprogramming genes and the lack of proven potential for differentiation into target cell types both in vitro and in vivo. Site-specific nucleases comprised of zinc finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs) and clustered regulated interspaced short palindromic repeats (CRISPRs) emerged as powerful genetic tools for precisely editing the genome, usurping the need for ESC-based genetic modifications even in the mouse. In this article, in the aftermath of these powerful genome editing technologies, the role of pluripotent stem cells in livestock genetics is discussed.

  19. Genetic modification of bone-marrow mesenchymal stem cells and hematopoietic cells with human coagulation factor IX-expressing plasmids.

    Science.gov (United States)

    Sam, Mohammad Reza; Azadbakhsh, Azadeh Sadat; Farokhi, Farrah; Rezazadeh, Kobra; Sam, Sohrab; Zomorodipour, Alireza; Haddad-Mashadrizeh, Aliakbar; Delirezh, Nowruz; Mokarizadeh, Aram

    2016-05-01

    Ex-vivo gene therapy of hemophilias requires suitable bioreactors for secretion of hFIX into the circulation and stem cells hold great potentials in this regard. Viral vectors are widely manipulated and used to transfer hFIX gene into stem cells. However, little attention has been paid to the manipulation of hFIX transgene itself. Concurrently, the efficacy of such a therapeutic approach depends on determination of which vectors give maximal transgene expression. With this in mind, TF-1 (primary hematopoietic lineage) and rat-bone marrow mesenchymal stem cells (BMSCs) were transfected with five hFIX-expressing plasmids containing different combinations of two human β-globin (hBG) introns inside the hFIX-cDNA and Kozak element and hFIX expression was evaluated by different methods. In BMSCs and TF-1 cells, the highest hFIX level was obtained from the intron-less and hBG intron-I,II containing plasmids respectively. The highest hFIX activity was obtained from the cells that carrying the hBG intron-I,II containing plasmids. BMSCs were able to produce higher hFIX by 1.4 to 4.7-fold increase with activity by 2.4 to 4.4-fold increase compared to TF-1 cells transfected with the same constructs. BMSCs and TF-1 cells could be effectively bioengineered without the use of viral vectors and hFIX minigene containing hBG introns could represent a particular interest in stem cell-based gene therapy of hemophilias.

  20. Modular projects and 'mean questions': best practices for advising an International Genetically Engineered Machines team.

    Science.gov (United States)

    Tsui, Jennifer; Meyer, Anne S

    2016-07-01

    In the yearly Internationally Genetically Engineered Machines (iGEM) competition, teams of Bachelor's and Master's students design and build an engineered biological system using DNA technologies. Advising an iGEM team poses unique challenges due to the inherent difficulties of mounting and completing a new biological project from scratch over the course of a single academic year; the challenges in obtaining financial and structural resources for a project that will likely not be fully realized; and conflicts between educational and competition-based goals. This article shares tips and best practices for iGEM team advisors, from two team advisors with very different experiences with the iGEM competition.

  1. Natural plant genetic engineer Agrobacterium rhizogenes: role of T-DNA in plant secondary metabolism.

    Science.gov (United States)

    Chandra, Sheela

    2012-03-01

    Agrobacterium rhizogenes is a natural plant genetic engineer. It is a gram-negative soil bacterium that induces hairy root formation. Success has been obtained in exploring the molecular mechanisms of transferred DNA (T-DNA) transfer, interaction with host plant proteins, plant defense signaling and integration to plant genome for successful plant genetic transformation. T-DNA and corresponding expression of rol genes alter morphology and plant host secondary metabolism. During transformation, there is a differential loss of a few T-DNA genes. Loss of a few ORFs drastically affect the growth and morphological patterns of hairy roots, expression pattern of biosynthetic pathway genes and accumulation of specific secondary metabolites.

  2. Differentiation of mesenchymal stem cells for cartilage tissue engineering: Individual and synergetic effects of three-dimensional environment and mechanical loading.

    Science.gov (United States)

    Panadero, J A; Lanceros-Mendez, S; Ribelles, J L Gomez

    2016-03-01

    Chondrogenesis of dedifferentiated chondrocytes and mesenchymal stem cells is influenced not only by soluble molecules like growth factors, but also by the cell environment itself. The latter is achieved through both mechanical cues - which act as stimulation factor and influences nutrient transport - and adhesion to extracellular matrix cues - which determine cell shape. Although the effects of soluble molecules and cell environment have been intensively addressed, few observations and conclusions about the interaction between the two have been achieved. In this work, we review the state of the art on the single effects between mechanical and biochemical cues, as well as on the combination of the two. Furthermore, we provide a discussion on the techniques currently used to determine the mechanical properties of materials and tissues generated in vitro, their limitations and the future research needs to properly address the identified problems. The importance of biomechanical cues in chondrogenesis is well known. This paper reviews the existing literature on the effect of mechanical stimulation on chondrogenic differentiation of mesenchymal stem cells in order to regenerate hyaline cartilage. Contradictory results found with respect to the effect of different modes of external loading can be explained by the different properties of the scaffolding system that holds the cells, which determine cell adhesion and morphology and spatial distribution of cells, as well as the stress transmission to the cells. Thus, this review seeks to provide an insight into the interplay between external loading program and scaffold properties during chondrogenic differentiation. The review of the literature reveals an important gap in the knowledge in this field and encourages new experimental studies. The main issue is that in each of the few cases in which the interplay is investigated, just two groups of scaffolds are compared, leaving intermediate adhesion conditions out of study

  3. The Discussions around Precision Genetic Engineering: Role of and Impact on Disabled People

    Directory of Open Access Journals (Sweden)

    Gregor Wolbring

    2016-09-01

    Full Text Available Genetic researchers are advancing in their abilities to extract precise genetic information from biological and human entities bringing genetic research steps closer to accurately modifying genes of biological entities, including that of humans. In this analytical essay, we focus on the discussions about precision genetic intervention that have taken place since March 2015 as they pertain to disabled people. We focus on two areas; one being the role of disabled people in the recent gene editing discussions and the second being the utility of existing legal instruments. Within our first focus we address the following questions: (a What is the visibility of disabled people in the gene-editing discussions that have taken place since March 2015? (b What has been the impact of those discussions on disabled people? (c Were social problems which disabled people face taken into account in those discussions; (d How does the reality of engagement with disabled people in these discussions fit with science, technology and innovation governance discourses that ask for more stakeholder, bottom up and anticipatory involvement? Within our second focus we address the following questions: (a What is the utility of the United Nations Convention on the Right of Persons with Disabilities (UNCRPD; and (b What is the utility of existing legal instruments covering genetic interventions: for preventing negative social consequences of genetic engineering developments for disabled people. We argue that (a the genetic engineering debates since March 2015 have portrayed disabled people dominantly through a medical lens; (b that the governance of science, technology and innovation of genetic engineering including anticipatory governance and responsible innovation discourses has not yet engaged with the social impact of gene editing on disabled people; (c that few scholars that focus on the social situation of disabled people are visible in the governance discussions of gene

  4. Role of nanotopography in the development of tissue engineered 3D organs and tissues using mesenchymal stem cells

    OpenAIRE

    Salmasi, Shima; Kalaskar, Deepak M.; Yoon, Wai-Weng; Blunn, Gordon W.; Seifalian, Alexander M

    2015-01-01

    Recent regenerative medicine and tissue engineering strategies (using cells, scaffolds, medical devices and gene therapy) have led to fascinating progress of translation of basic research towards clinical applications. In the past decade, great deal of research has focused on developing various three dimensional (3D) organs, such as bone, skin, liver, kidney and ear, using such strategies in order to replace or regenerate damaged organs for the purpose of maintaining or restoring organs’ func...

  5. Genetic engineering of flavonoid pigments to modify flower color in floricultural plants.

    Science.gov (United States)

    Nishihara, Masahiro; Nakatsuka, Takashi

    2011-03-01

    Recent advances in genetic transformation techniques enable the production of desirable and novel flower colors in some important floricultural plants. Genetic engineering of novel flower colors is now a practical technology as typified by commercialization of a transgenic blue rose and blue carnation. Many researchers exploit knowledge of flavonoid biosynthesis effectively to obtain unique flower colors. So far, the main pigments targeted for flower color modification are anthocyanins that contribute to a variety of colors such as red, pink and blue, but recent studies have also utilized colorless or faint-colored compounds. For example, chalcones and aurones have been successfully engineered to produce yellow flowers, and flavones and flavonols used to change flower color hues. In this review, we summarize examples of successful flower color modification in floricultural plants focusing on recent advances in techniques.

  6. Application of genetically engineered microbial whole-cell biosensors for combined chemosensing.

    Science.gov (United States)

    He, Wei; Yuan, Sheng; Zhong, Wen-Hui; Siddikee, Md Ashaduzzaman; Dai, Chuan-Chao

    2016-02-01

    The progress of genetically engineered microbial whole-cell biosensors for chemosensing and monitoring has been developed in the last 20 years. Those biosensors respond to target chemicals and produce output signals, which offer a simple and alternative way of assessment approaches. As actual pollution caused by human activities usually contains a combination of different chemical substances, how to employ those biosensors to accurately detect real contaminant samples and evaluate biological effects of the combined chemicals has become a realistic object of environmental researches. In this review, we outlined different types of the recent method of genetically engineered microbial whole-cell biosensors for combined chemical evaluation, epitomized their detection performance, threshold, specificity, and application progress that have been achieved up to now. We also discussed the applicability and limitations of this biosensor technology and analyzed the optimum conditions for their environmental assessment in a combined way.

  7. Genetic Algorithm and Fuzzy Tuning PID Controller Applied on Speed Control System for Marine Diesel Engines

    Directory of Open Access Journals (Sweden)

    Naeim Farouk

    2012-11-01

    Full Text Available The degree of speed control of ship machinery effects on the economics and optimization of the machinery configuration and operation. All marine vessel ranging need some sort of speed control system to control and govern the speed of the marine diesel engines. The main focus of this study is to apply and comparative between two specific soft-computing techniques. Fuzzy logic controller and genetic algorithm to design and tuning of PID controller for applied on speed control system of marine diesel engine to get an output with better dynamic and static performance. Simulation results show that the response of system when using genetic algorithm is better and faster than when using fuzzy tuning PID controller.

  8. Nuclear and plastid genetic engineering of plants: comparison of opportunities and challenges.

    Science.gov (United States)

    Meyers, Benjamin; Zaltsman, Adi; Lacroix, Benoît; Kozlovsky, Stanislav V; Krichevsky, Alexander

    2010-01-01

    Plant genetic engineering is one of the key technologies for crop improvement as well as an emerging approach for producing recombinant proteins in plants. Both plant nuclear and plastid genomes can be genetically modified, yet fundamental functional differences between the eukaryotic genome of the plant cell nucleus and the prokaryotic-like genome of the plastid will have an impact on key characteristics of the resulting transgenic organism. So, which genome, nuclear or plastid, to transform for the desired transgenic phenotype? In this review we compare the advantages and drawbacks of engineering plant nuclear and plastid genomes to generate transgenic plants with the traits of interest, and evaluate the pros and cons of their use for different biotechnology and basic research applications, ranging from generation of commercial crops with valuable new phenotypes to 'bioreactor' plants for large-scale production of recombinant proteins to research model plants expressing various reporter proteins.

  9. Genetic And Metabolic Engineering Of Microorganisms For The Development Of New Flavor Compounds From Terpenic Substrates

    OpenAIRE

    Bution; Murillo L.; Molina; Gustavo; Abrahao; Meissa R. E.; Pastore; Glaucia M.

    2015-01-01

    Throughout human history, natural products have been the basis for the discovery and development of therapeutics, cosmetic and food compounds used in industry. Many compounds found in natural organisms are rather difficult to chemically synthesize and to extract in large amounts, and in this respect, genetic and metabolic engineering are playing an increasingly important role in the production of these compounds, such as new terpenes and terpenoids, which may potentially be used to create aro...

  10. Recent advances in plant biotechnology and genetic engineering for production of secondary metabolites.

    Science.gov (United States)

    Sheludko, Y V

    2010-01-01

    For a long time people are using plants not only as crop cultures but also for obtaining of various chemicals. Currently plants remain one of the most important and essential sources of biologically active compounds in spite of progress in chemical or microbial synthesis. In our review we compare potentials and perspectives of modern genetic engineering approaches for pharmaceutical biotechnology and give examples of actual biotechnological systems used for production of several promising natural compounds: artemisinin, paclitaxel and scopolamine.

  11. Overview of KRAS-Driven Genetically Engineered Mouse Models of Non-Small Cell Lung Cancer.

    Science.gov (United States)

    Sheridan, Clare; Downward, Julian

    2015-01-01

    KRAS, the most frequently mutated oncogene in non-small cell lung cancer, has been utilized extensively to model human lung adenocarcinomas. The results from such studies have enhanced considerably an understanding of the relationship between KRAS and the development of lung cancer. Detailed in this overview are the features of various KRAS-driven genetically engineered mouse models (GEMMs) of non-small cell lung cancer, their utilization, and the potential of these models for the study of lung cancer biology.

  12. [Genetic engineering technologies of stimulating angiogenesis as an innovation trend in angiology and vascular surgery].

    Science.gov (United States)

    Gavrilenko, A V; Voronov, D A

    2015-01-01

    Presented herein is a review of the principles, fundamental concepts, and possibilities of genetic engineering technologies of stimulating angiogenesis for treatment of patients with lower limb chronic ischaemia. This is followed by a detailed discussion of the structure and results of Russian and foreign studies on this direction, also considering the causes of differences of their results. Outlined is a circle of clinical situations in relation to which these technologies may be regarded as most promising.

  13. Growth of genetically engineered Pseudomonas aeruginosa and Pseudomonas putida in soil and rhizosphere.

    OpenAIRE

    Yeung, K H; Schell, M A; Hartel, P G

    1989-01-01

    The effect of the addition of a recombinant plasmid containing the pglA gene encoding an alpha-1,4-endopolygalacturonase from Pseudomonas solanacearum on the growth of Pseudomonas aeruginosa and Pseudomonas putida in soil and rhizosphere was determined. Despite a high level of polygalacturonase production by genetically engineered P. putida and P. aeruginosa, the results suggest that polygalacturonase production had little effect on the growth of these strains in soil or rhizosphere.

  14. [Genetic engineering and assisted reproduction techniques in man: a framework for sociologic analysis].

    Science.gov (United States)

    Sánchez Morales, M R

    1999-01-01

    The possibilities opened up by genetic engineering and assisted reproduction techniques require reflection by sociologists and extensive public debate. In view of their potential as factors of social change, evaluation and control are warranted. They can be viable only if transparent and through public co-responsibility, for which an exchange of views is needed between all those who play a part in the development of said techniques. This dialogue must be wholly interdisciplinary and democratic.

  15. An effective hybrid cuckoo search and genetic algorithm for constrained engineering design optimization

    Science.gov (United States)

    Kanagaraj, G.; Ponnambalam, S. G.; Jawahar, N.; Mukund Nilakantan, J.

    2014-10-01

    This article presents an effective hybrid cuckoo search and genetic algorithm (HCSGA) for solving engineering design optimization problems involving problem-specific constraints and mixed variables such as integer, discrete and continuous variables. The proposed algorithm, HCSGA, is first applied to 13 standard benchmark constrained optimization functions and subsequently used to solve three well-known design problems reported in the literature. The numerical results obtained by HCSGA show competitive performance with respect to recent algorithms for constrained design optimization problems.

  16. In-depth metabolic phenotyping of genetically engineered mouse models in obesity and diabetes.

    Science.gov (United States)

    Lee, Hui-Young; Jeong, Kyeong-Hoon; Choi, Cheol Soo

    2014-10-01

    The world-wide prevalence of obesity and diabetes has increased sharply during the last two decades. Accordingly, the metabolic phenotyping of genetically engineered mouse models is critical for evaluating the functional roles of target genes in obesity and diabetes, and for developing new therapeutic targets. In this review, we discuss the practical meaning of metabolic phenotyping, the strategy of choosing appropriate tests, and considerations when designing and performing metabolic phenotyping in mice.

  17. Telos, conservation of welfare, and ethical issues in genetic engineering of animals.

    Science.gov (United States)

    Rollin, Bernard E

    2015-01-01

    The most long-lived metaphysics or view of reality in the history of Western thought is Aristotle's teleology, which reigned for almost 2,000 years. Biology was expressed in terms of function or telos, and accorded perfectly with common sense. The rise of mechanistic, Newtonian science vanquished teleological explanations. Understanding and accommodating animal telos was essential to success in animal husbandry, which involved respect for telos, and was presuppositional to our "ancient contract" with domestic animals. Telos was further abandoned with the rise of industrial agriculture, which utilized "technological fixes" to force animal into environments they were unsuited for, while continuing to be productive. Loss of husbandry and respect for telos created major issues for farm animal welfare, and forced the creation of a new ethic demanding respect for telos. As genetic engineering developed, the notion arose of modifying animals to fit their environment in order to avoid animal suffering, rather than fitting them into congenial environments. Most people do not favor changing the animals, rather than changing the conditions under which they are reared. Aesthetic appreciation of husbandry and virtue ethics militate in favor of restoring husbandry, rather than radically changing animal teloi. One, however, does not morally wrong teloi by changing them-one can only wrong individuals. In biomedical research, we do indeed inflict major pain, suffering and disease on animals. And genetic engineering seems to augment our ability to create animals to model diseases, particularly more than 3,000 known human genetic diseases. The disease, known as Lesch-Nyhan's syndrome or HPRT deficiency, which causes self-mutilation and mental retardation, provides us with a real possibility for genetically creating "animal models" of this disease, animals doomed to a life of great and unalleviable suffering. This of course creates a major moral dilemma. Perhaps one can use the very

  18. Preparation and properties of microencapsulated genetically engineered bacteria cells for oral therapy of uremia

    Institute of Scientific and Technical Information of China (English)

    GAO Hong; YU Yaoting; CAI Baoli; WANG Manyan

    2004-01-01

    Microencapsulated genetically engineered bacteria cells are a novel approach of oral therapy for uremia.Klebsiella aerogenes urease genes (UreaDABCEFG) are transformed into E. coli DH5α cells through plasmid pKAU17. The transformant can use urea or ammonia as its sole nitrogen source through strain training. The urease genetically engineered bacteria cells are entrapped in polyvinyl alcohol (PVA) microcapsules, which can be used to remove urea from uremia patients. The mechanical strength of PVA microcapsules is significantly higher than that of APA microcapsules. This suggests that the problem of friability of APA can be solved in this way. The optimal conditions for the preparation of PVA microencapsulated genetically engineered bacterial cells are: polyvinyl alcohol (PVA, 2450±50)used as the carrier at a concentration 6%, the pH value of boric acid as crosslinking reagent 6.5, crosslinking time 24 h,entrapment ratio of bacteria 8%, air flow rate of the encapsulate device 3 L/min and liquid flow rate at 1 mL/10 min.The average diameter of microcapsules prepared under these optimal conditions is 20-40 mesh. Experiments in vitro showed that one hundred milligrams of wet bacterial cells in PVA microcapsules could remove 18.4 mg of urea in 4 h.

  19. Genetic engineering of mice to test the oxidative damage theory of aging.

    Science.gov (United States)

    Martin, George M

    2005-12-01

    The laboratory mouse Mus musculus domesticus provides the best current mammalian models for the genetic analysis of aging. We give a brief overview of the use of transgenic manipulations to test the oxidative damage theory of aging. These manipulations are of two types: The first approach engineers mice that exhibit increased sensitivities to oxidative damage and thus produces mice that are likely to be short-lived. The second approach engineers mice to be more resistant to such injuries, and thus may produce mice that exhibit enhanced longevities, something that is much harder to engineer. The latter result is thus more meaningful, with the caveat that it may result from some special vulnerability of a particular lab strain or lab strains in general. The first approach, most elegantly carried out by Arlan Richardson's laboratory, provides evidence against the oxidative damage theory. My colleagues and I have been engaged in the second approach and have accumulated evidence supporting the theory. These conventional transgenic experiments, however, should be supplemented by alternative genetic approaches. One that is surprisingly neglected takes advantage of the pleuripotency of embryonic stem cells and the power of somatic cell genetics. A cautionary note is that interventions that minimize oxidative stress may be complicated by unwanted compromises of physiologically adaptive actions such as superoxide signaling and the possible protective effects of certain oxidatively modified proteins.

  20. Current status of genetic engineering in cotton (Gossypium hirsutum L): an assessment.

    Science.gov (United States)

    Chakravarthy, Vajhala S K; Reddy, Tummala Papi; Reddy, Vudem Dashavantha; Rao, Khareedu Venkateswara

    2014-06-01

    Cotton is considered as the foremost commercially important fiber crop and is deemed as the backbone of the textile industry. The productivity of cotton crop, worldwide, is severely hampered by the occurrence of pests, weeds, pathogens apart from various environmental factors. Several beneficial agronomic traits, viz., early maturity, improved fiber quality, heat tolerance, etc. have been successfully incorporated into cotton varieties employing conventional hybridization and mutation breeding. Crop losses, due to biotic factors, are substantial and may be reduced through certain crop protection strategies. In recent years, pioneering success has been achieved through the adoption of modern biotechnological approaches. Genetically engineered cotton varieties, expressing Bacillus thuringiensis cry genes, proved to be highly successful in controlling the bollworm complex. Various other candidate genes responsible for resistance to insect pests and pathogens, tolerance to major abiotic stress factors such as temperature, drought and salinity, have been introduced into cotton via genetic engineering methods to enhance the agronomic performance of cotton cultivars. Furthermore, genes for improving the seed oil quality and fiber characteristics have been identified and introduced into cotton cultivars. This review provides a brief overview of the various advancements made in cotton through genetic engineering approaches.

  1. Controlled Dual Growth Factor Delivery From Microparticles Incorporated Within Human Bone Marrow-Derived Mesenchymal Stem Cell Aggregates for Enhanced Bone Tissue Engineering via Endochondral Ossification.

    Science.gov (United States)

    Dang, Phuong N; Dwivedi, Neha; Phillips, Lauren M; Yu, Xiaohua; Herberg, Samuel; Bowerman, Caitlin; Solorio, Loran D; Murphy, William L; Alsberg, Eben

    2016-02-01

    Bone tissue engineering via endochondral ossification has been explored by chondrogenically priming cells using soluble mediators for at least 3 weeks to produce a hypertrophic cartilage template. Although recapitulation of endochondral ossification has been achieved, long-term in vitro culture is required for priming cells through repeated supplementation of inductive factors in the media. To address this challenge, a microparticle-based growth factor delivery system was engineered to drive endochondral ossification within human bone marrow-derived mesenchymal stem cell (hMSC) aggregates. Sequential exogenous presentation of soluble transforming growth factor-β1 (TGF-β1) and bone morphogenetic protein-2 (BMP-2) at various defined time courses resulted in varying degrees of chondrogenesis and osteogenesis as demonstrated by glycosaminoglycan and calcium content. The time course that best induced endochondral ossification was used to guide the development of the microparticle-based controlled delivery system for TGF-β1 and BMP-2. Gelatin microparticles capable of relatively rapid release of TGF-β1 and mineral-coated hydroxyapatite microparticles permitting more sustained release of BMP-2 were then incorporated within hMSC aggregates and cultured for 5 weeks following the predetermined time course for sequential presentation of bioactive signals. Compared with cell-only aggregates treated with exogenous growth factors, aggregates with incorporated TGF-β1- and BMP-2-loaded microparticles exhibited enhanced chondrogenesis and alkaline phosphatase activity at week 2 and a greater degree of mineralization by week 5. Staining for types I and II collagen, osteopontin, and osteocalcin revealed the presence of cartilage and bone. This microparticle-incorporated system has potential as a readily implantable therapy for healing bone defects without the need for long-term in vitro chondrogenic priming. Significance: This study demonstrates the regulation of chondrogenesis

  2. Isolation and trans-differentiation of mesenchymal stromal cells into smooth muscle cells: Utility and applicability for cell-sheet engineering.

    Science.gov (United States)

    Shudo, Yasuhiro; Cohen, Jeffrey E; Goldstone, Andrew B; MacArthur, John W; Patel, Jay; Edwards, Bryan B; Hopkins, Michael S; Steele, Amanda N; Joubert, Lydia-Marie; Miyagawa, Shigeru; Sawa, Yoshiki; Woo, Y Joseph

    2016-04-01

    Bone marrow (BM)-derived mesenchymal stromal cells (MSCs) have shown potential to differentiate into various cell types, including smooth muscle cells (SMCs). The extracellular matrix (ECM) represents an appealing and readily available source of SMCs for use in tissue engineering. In this study, we hypothesized that the ECM could be used to induce MSC differentiation to SMCs for engineered cell-sheet construction. Primary MSCs were isolated from the BM of Wistar rats, transferred and cultured on dishes coated with 3 different types of ECM: collagen type IV (Col IV), fibronectin (FN), and laminin (LM). Primary MSCs were also included as a control. The proportions of SMC (a smooth muscle actin [aSMA] and SM22a) and MSC markers were examined with flow cytometry and Western blotting, and cell proliferation rates were also quantified. Both FN and LM groups were able to induce differentiation of MSCs toward smooth muscle-like cell types, as evidenced by an increase in the proportion of SMC markers (aSMA; Col IV 42.3 ± 6.9%, FN 65.1 ± 6.5%, LM 59.3 ± 7.0%, Control 39.9 ± 3.1%; P = 0.02, SM22; Col IV 56.0 ± 7.7%, FN 74.2 ± 6.7%, LM 60.4 ± 8.7%, Control 44.9 ± 3.6%) and a decrease in that of MSC markers (CD105: Col IV 64.0 ± 5.2%, FN 57.6 ± 4.0%, LM 60.3 ± 7.0%, Control 85.3 ± 4.2%; P = 0.03). The LM group showed a decrease in overall cell proliferation, whereas FN and Col IV groups remained similar to control MSCs (Col IV, 9.0 ± 2.3%; FN, 9.8 ± 2.5%; LM, 4.3 ± 1.3%; Control, 9.8 ± 2.8%). Our findings indicate that ECM selection can guide differentiation of MSCs into the SMC lineage. Fibronectin preserved cellular proliferative capacity while yielding the highest proportion of differentiated SMCs, suggesting that FN-coated materials may be facilitate smooth muscle tissue engineering. Copyright © 2016 International Society for Cellular Therapy. Published by Elsevier Inc. All rights

  3. A CRISPR-Cas9 System for Genetic Engineering of Filamentous Fungi.

    Science.gov (United States)

    Nødvig, Christina S; Nielsen, Jakob B; Kogle, Martin E; Mortensen, Uffe H

    2015-01-01

    The number of fully sequenced fungal genomes is rapidly increasing. Since genetic tools are poorly developed for most filamentous fungi, it is currently difficult to employ genetic engineering for understanding the biology of these fungi and to fully exploit them industrially. For that reason there is a demand for developing versatile methods that can be used to genetically manipulate non-model filamentous fungi. To facilitate this, we have developed a CRISPR-Cas9 based system adapted for use in filamentous fungi. The system is simple and versatile, as RNA guided mutagenesis can be achieved by transforming a target fungus with a single plasmid. The system currently contains four CRISPR-Cas9 vectors, which are equipped with commonly used fungal markers allowing for selection in a broad range of fungi. Moreover, we have developed a script that allows identification of protospacers that target gene homologs in multiple species to facilitate introduction of common mutations in different filamentous fungi. With these tools we have performed RNA-guided mutagenesis in six species of which one has not previously been genetically engineered. Moreover, for a wild-type Aspergillus aculeatus strain, we have used our CRISPR Cas9 system to generate a strain that contains an AACU_pyrG marker and demonstrated that the resulting strain can be used for iterative gene targeting.

  4. The potential of 3-dimensional construct engineered from poly(lactic-co-glycolic acid)/fibrin hybrid scaffold seeded with bone marrow mesenchymal stem cells for in vitro cartilage tissue engineering.

    Science.gov (United States)

    Abdul Rahman, Rozlin; Mohamad Sukri, Norhamiza; Md Nazir, Noorhidayah; Ahmad Radzi, Muhammad Aa'zamuddin; Zulkifly, Ahmad Hafiz; Che Ahmad, Aminudin; Hashi, Abdurezak Abdulahi; Abdul Rahman, Suzanah; Sha'ban, Munirah

    2015-08-01

    Articular cartilage is well known for its simple uniqueness of avascular and aneural structure that has limited capacity to heal itself when injured. The use of three dimensional construct in tissue engineering holds great potential in regenerating cartilage defects. This study evaluated the in vitro cartilaginous tissue formation using rabbit's bone marrow mesenchymal stem cells (BMSCs)-seeded onto poly(lactic-co-glycolic acid) PLGA/fibrin and PLGA scaffolds. The in vitro cartilaginous engineered constructs were evaluated by gross inspection, histology, cell proliferation, gene expression and sulphated glycosaminoglycan (sGAG) production at week 1, 2 and 3. After 3 weeks of culture, the PLGA/fibrin construct demonstrated gross features similar to the native tissue with smooth, firm and glistening appearance, superior histoarchitectural and better cartilaginous extracellular matrix compound in concert with the positive glycosaminoglycan accumulation on Alcian blue. Significantly higher cell proliferation in PLGA/fibrin construct was noted at day-7, day-14 and day-21 (p<0.05 respectively). Both constructs expressed the accumulation of collagen type II, collagen type IX, aggrecan and sox9, showed down-regulation of collagen type I as well as produced relative sGAG content with PLGA/fibrin construct exhibited better gene expression in all profiles and showed significantly higher relative sGAG content at each time point (p<0.05). This study suggested that with optimum in vitro manipulation, PLGA/fibrin when seeded with pluripotent non-committed BMSCs has the capability to differentiate into chondrogenic lineage and may serve as a prospective construct to be developed as functional tissue engineered cartilage.

  5. Reproductive cloning, genetic engineering and the autonomy of the child: the moral agent and the open future.

    Science.gov (United States)

    Mameli, M

    2007-02-01

    Some authors have argued that the human use of reproductive cloning and genetic engineering should be prohibited because these biotechnologies would undermine the autonomy of the resulting child. In this paper, two versions of this view are discussed. According to the first version, the autonomy of cloned and genetically engineered people would be undermined because knowledge of the method by which these people have been conceived would make them unable to assume full responsibility for their actions. According to the second version, these biotechnologies would undermine autonomy by violating these people's right to an open future. There is no evidence to show that people conceived through cloning and genetic engineering would inevitably or even in general be unable to assume responsibility for their actions; there is also no evidence for the claim that cloning and genetic engineering would inevitably or even in general rob the child of the possibility to choose from a sufficiently large array of life plans.

  6. A CAL Program to Teach the Basic Principles of Genetic Engineering--A Change from the Traditional Approach.

    Science.gov (United States)

    Dewhurst, D. G.; And Others

    1989-01-01

    An interactive computer-assisted learning program written for the BBC microcomputer to teach the basic principles of genetic engineering is described. Discussed are the hardware requirements software, use of the program, and assessment. (Author/CW)

  7. A CAL Program to Teach the Basic Principles of Genetic Engineering--A Change from the Traditional Approach.

    Science.gov (United States)

    Dewhurst, D. G.; And Others

    1989-01-01

    An interactive computer-assisted learning program written for the BBC microcomputer to teach the basic principles of genetic engineering is described. Discussed are the hardware requirements software, use of the program, and assessment. (Author/CW)

  8. Enhanced osteogenesis of human alveolar bone-derived mesenchymal stem cells for tooth tissue engineering using fluid shear stress in a rocking culture method.

    Science.gov (United States)

    Lim, Ki-Taek; Kim, Jangho; Seonwoo, Hoon; Chang, Jung Uk; Choi, Hwajung; Hexiu, Jin; Cho, Woo Jae; Choung, Pill-Hoon; Chung, Jong Hoon

    2013-02-01

    This study instituted a simple approach to stimulate alveolar bone regeneration for tooth tissue engineering by controlling effects of low fluid dynamic shear stress (LFDSS) on growth and differentiation in vitro. Human alveolar bone-derived mesenchymal stem cells (hABMSCs) harvested from human mandibular alveolar bone were cultured with LFDSS to generate cultures containing bone-like formations. To distinguish between osteodifferentiation and bone-like formation, cells were cultured either with or without fluid shear stress. The calcium content and alkaline phosphatase (ALP) activity of hABMSCs were used as indicators of osteogenesis. Cell viability and proliferation after stimulating with LFDSS for 10-60 min/day were higher than with longer stimulations. Mineralized nodules formed when osteoblasts were cultured with an induction medium, a marker of osteogenic differentiation. ALP activity tended to increase after 10 and 60 min/day of stimulation. In addition, LFDSS conditions also increased gene expression of IBSP, RUNX2, COL-I, ALP, OCN, and OPN, as shown by reverse transcriptase-polymerase chain reaction. From the results of a proteomics array, LFDSS groups were intensely expressed with several factors (EGF, HGF, IGF, TGF, and PDGF). Furthermore, CD146 and Stro-1 expression increased in cells treated with 30 min/day and decreased in cells treated with 120 min/day, as determined by cell surface antigen analysis by fluorescence-activated cell-sorting analysis. These results strongly showed that LFDSS at the proper intensity and time enhanced the differentiation and maturation of hABMSCs. In conclusion, an appropriate level of LFDSS can potently and positively modulate proliferation and differentiation in hABMSCs.

  9. Ectopic osteogenesis and scaffold biodegradation of tissue engineering bone composed of chitosan and osteo-induced bone marrow mesenchymal stem cells in vivo

    Institute of Scientific and Technical Information of China (English)

    He Yiqun; Dong Youhai; Chen Xujun; Lin Rongqiang

    2014-01-01

    Background Chitosan (CS) scaffolds combined with osteogenically induced bone marrow mesenchymal stem cells (BMSCs) have been proved to be promising substitutes for repairing bone defects.Nevertheless,the bone-forming and scaffold-biodegrading processes are seldom studied.This study aimed to determine the osteogenic ability of CS/osteoinduced BMSC composites by observing the bone-forming process and explore the relationship between bone formation and scaffold biodegradation.Methods The CS/osteo-induced BMSC composites (CS+cells group) and the CS scaffolds (CS group) were,respectively,implanted into SD rat thigh muscles.At 2,4,6,8,and 12 weeks postoperatively,the rat femurs were scanned by CT,and the CT values of the implants were measured and comparatively analyzed.Subsequently,the implants were harvested and stained with hematoxylin and eosin and Masson trichrome,and the percentages of bone area,scaffold area,and collagen area were calculated and compared between the two groups.Results The imaging results showed that the densities of implants of the two groups gradually increased along with time,but the CT values of implants in the CS+cells group were much higher than in the CS group at the same time point (P <0.05).The histological results showed that the de novo bone and collagen formed in the pores of the scaffolds and gradually increased since 2 weeks postoperation in both groups,and the scaffold gradually degraded along with the boneforming process.However,the comparative analysis results showed that the CS+cells group gained more de novo bone and collagen formation and had less scaffold than the CS group at the same time point (P <0.05).Conclusion The CS/osteo-induced BMSC composites are excellent bone tissue engineering substitutes,and the scaffold biodegradation is accordant with the bone formation.

  10. Del-1 Overexpression in Endothelial Cells Increases Vascular Density in Tissue-Engineered Implants Containing Endothelial Cells and Adipose-Derived Mesenchymal Stromal Cells

    Science.gov (United States)

    Ciucurel, Ema C.

    2014-01-01

    We used a combination of strategies to stimulate the vascularization of tissue-engineered constructs in vivo including a modular approach to build larger tissues from individual building blocks (“modules”) mixed together. Each building block included vascular cells by design; modules were submillimeter-sized collagen gels with an outer layer of endothelial cells (ECs), and with embedded adipose-derived mesenchymal stromal cells (adMSCs) to support EC survival and blood vessel maturation in vivo. We transduced the ECs that coat the modules with a lentiviral construct to overexpress the angiogenic extracellular matrix (ECM) protein Developmental endothelial locus-1 (Del-1). Upon injection of modules in a subcutaneous SCID/Bg mouse model, there was an increase in the number of blood vessels for implants with ECs transduced to overexpress Del-1 compared with control implants (with enhanced green fluorescent protein [eGFP]–transduced ECs) over the 21-day duration of the study. The greatest difference between Del-1 and eGFP implants and the highest number of blood vessels were observed 7 days after transplantation. The day-7 Del-1 implants also had increased SMA+ staining compared with control, suggesting increased blood vessel maturation through recruitment of SMA+ smooth muscle cells or pericytes to stabilize the newly formed blood vessels. Perfusion studies (microcomputed tomography, ultrasound imaging, and systemic injection of fluorescent UEA-1 or dextran) showed that some of the newly formed blood vessels (both donor derived and host derived, in both Del-1 and eGFP implants) were perfused and connected to the host vasculature as early as 7 days after transplantation, and at later time points as well. Nevertheless, perfusion of the implants was limited in some cases, suggesting that further improvements are necessary to normalize the vasculature at the implant site. PMID:24151812

  11. Research Survey of Genetic Engineering Drugs%基因工程药物研究概况

    Institute of Scientific and Technical Information of China (English)

    郭俊清; 徐进; 李建正

    2011-01-01

    对新发展起来的产业基因工程药物的研究作了简要的概述,通过对其发展历史及当前的几种药物的叙述,预测其发展前景。%The new genetically engineered drug industry research was summarized briefly. The prospect of genetically engineered drug industry research was predicted by describing its developing history and several current drugs.

  12. Testing Current and Developing Novel Therapies for NF1-Mutant Sarcomas in a Genetically Engineered Mouse Model

    Science.gov (United States)

    2015-04-01

    1   AWARD NUMBER: W81XWH-14-1-0067 TITLE: Testing Current and Developing Novel Therapies for NF1 -Mutant Sarcomas in a Genetically Engineered...Mar 2014 - 14 Mar 2015 4. TITLE AND SUBTITLE Testing Current and Developing Novel Therapies for NF1 - Mutant Sarcomas in a Genetically Engineered...Patients with Neurofibromatosis type 1 ( NF1 ) are at increased risk for developing malignant tumors of the connective tissue called soft-tissue sarcomas

  13. Degradation of phenanthrene and pyrene using genetically engineered dioxygenase producing Pseudomonas putida in soil

    Directory of Open Access Journals (Sweden)

    Mardani Gashtasb

    2016-01-01

    Full Text Available Bioremediation use to promote degradation and/or removal of contaminants into nonhazardous or less-hazardous substances from the environment using microbial metabolic ability. Pseudomonas spp. is one of saprotrophic soil bacterium and can be used for biodegradation of polycyclic aromatic hydrocarbons (PAHs but this activity in most species is weak. Phenanthrene and pyrene could associate with a risk of human cancer development in exposed individuals. The aim of the present study was application of genetically engineered P. putida that produce dioxygenase for degradation of phenanthrene and pyrene in spiked soil using high-performance liquid chromatography (HPLC method. The nahH gene that encoded catechol 2,3-dioxygenase (C23O was cloned into pUC18 and pUC18-nahH recombinant vector was generated and transformed into wild P. putida, successfully. The genetically modified and wild types of P. putida were inoculated in soil and pilot plan was prepared. Finally, degradation of phenanthrene and pyrene by this bacterium in spiked soil were evaluated using HPLC measurement technique. The results were showed elimination of these PAH compounds in spiked soil by engineered P. putida comparing to dishes containing natural soil with normal microbial flora and inoculated autoclaved soil by wild type of P. putida were statistically significant (p0.05 but it was few impact on this process (more than 2%. Additional and verification tests including catalase, oxidase and PCR on isolated bacteria from spiked soil were indicated that engineered P. putida was alive and functional as well as it can affect on phenanthrene and pyrene degradation via nahH gene producing. These findings indicated that genetically engineered P. putida generated in this work via producing C23O enzyme can useful and practical for biodegradation of phenanthrene and pyrene as well as petroleum compounds in polluted environments.

  14. Open field release of genetically engineered sterile male Aedes aegypti in Malaysia.

    Directory of Open Access Journals (Sweden)

    Renaud Lacroix

    Full Text Available BACKGROUND: Dengue is the most important mosquito-borne viral disease. In the absence of specific drugs or vaccines, control focuses on suppressing the principal mosquito vector, Aedes aegypti, yet current methods have not proven adequate to control the disease. New methods are therefore urgently needed, for example genetics-based sterile-male-release methods. However, this requires that lab-reared, modified mosquitoes be able to survive and disperse adequately in the field. METHODOLOGY/PRINCIPAL FINDINGS: Adult male mosquitoes were released into an uninhabited forested area of Pahang, Malaysia. Their survival and dispersal was assessed by use of a network of traps. Two strains were used, an engineered 'genetically sterile' (OX513A and a wild-type laboratory strain, to give both absolute and relative data about the performance of the modified mosquitoes. The two strains had similar maximum dispersal distances (220 m, but mean distance travelled of the OX513A strain was lower (52 vs. 100 m. Life expectancy was similar (2.0 vs. 2.2 days. Recapture rates were high for both strains, possibly because of the uninhabited nature of the site. CONCLUSIONS/SIGNIFICANCE: After extensive contained studies and regulatory scrutiny, a field release of engineered mosquitoes was safely and successfully conducted in Malaysia. The engineered strain showed similar field longevity to an unmodified counterpart, though in this setting dispersal was reduced relative to the unmodified strain. These data are encouraging for the future testing and implementation of genetic control strategies and will help guide future field use of this and other engineered strains.

  15. An improved ARS2-derived nuclear reporter enhances the efficiency and ease of genetic engineering in Chlamydomonas

    DEFF Research Database (Denmark)

    Specht, Elizabeth A; Nour-Eldin, Hussam Hassan; Hoang, Kevin T D

    2015-01-01

    The model alga Chlamydomonas reinhardtii has been used to pioneer genetic engineering techniques for high-value protein and biofuel production from algae. To date, most studies of transgenic Chlamydomonas have utilized the chloroplast genome due to its ease of engineering, with a sizeable suite...

  16. Ray Wu,Cornell’s acclaimed pioneer of genetic engineering and developer of insect-resistant rice

    Institute of Scientific and Technical Information of China (English)

    2009-01-01

    ITHACA, N.Y. -- Ray J. Wu, Cornell University professor of molecular biology and genetics, who was widely recog-nized as one of the fathers of genetic engineering and who developed and sought to feed the world with a higher yield-ing rice that resists insects and drought, died of cardiac arrest in Ithaca, Feb. 10.

  17. Effect of synthetic auxin herbicides on seed development and viability in genetically-engineered glyphosate-resistant alfalfa

    Science.gov (United States)

    Feral populations of cultivated crops have the potential to function as bridges and reservoirs that contribute to the unwanted movement of novel genetically engineered (GE) traits. Recognizing that feral alfalfa has the potential to lower genetic purity in alfalfa seed production fields when it is g...

  18. Genetic and metabolic engineering of microorganisms for the development of new flavor compounds from terpenic substrates.

    Science.gov (United States)

    Bution, Murillo L; Molina, Gustavo; Abrahão, Meissa R E; Pastore, Gláucia M

    2015-01-01

    Throughout human history, natural products have been the basis for the discovery and development of therapeutics, cosmetic and food compounds used in industry. Many compounds found in natural organisms are rather difficult to chemically synthesize and to extract in large amounts, and in this respect, genetic and metabolic engineering are playing an increasingly important role in the production of these compounds, such as new terpenes and terpenoids, which may potentially be used to create aromas in industry. Terpenes belong to the largest class of natural compounds, are produced by all living organisms and play a fundamental role in human nutrition, cosmetics and medicine. Recent advances in systems biology and synthetic biology are allowing us to perform metabolic engineering at the whole-cell level, thus enabling the optimal design of microorganisms for the efficient production of drugs, cosmetic and food additives. This review describes the recent advances made in the genetic and metabolic engineering of the terpenes pathway with a particular focus on systems biotechnology.

  19. Genetically engineering cyanobacteria to convert CO₂, water, and light into the long-chain hydrocarbon farnesene.

    Science.gov (United States)

    Halfmann, Charles; Gu, Liping; Gibbons, William; Zhou, Ruanbao

    2014-12-01

    Genetically engineered cyanobacteria offer a shortcut to convert CO2 and H2O directly into biofuels and high value chemicals for societal benefits. Farnesene, a long-chained hydrocarbon (C15H24), has many applications in lubricants, cosmetics, fragrances, and biofuels. However, a method for the sustainable, photosynthetic production of farnesene has been lacking. Here, we report the photosynthetic production of farnesene by the filamentous cyanobacterium Anabaena sp. PCC 7120 using only CO2, mineralized water, and light. A codon-optimized farnesene synthase gene was chemically synthesized and then expressed in the cyanobacterium, enabling it to synthesize farnesene through its endogenous non-mevalonate (MEP) pathway. Farnesene excreted from the engineered cyanobacterium volatilized into the flask head space and was recovered by adsorption in a resin column. The maximum photosynthetic productivity of farnesene was 69.1 ± 1.8 μg·L(-1)·O.D.(-1)·d(-1). Compared to the wild type, the farnesene-producing cyanobacterium also exhibited a 60 % higher PSII activity under high light, suggesting increased farnesene productivity in such conditions. We envision genetically engineered cyanobacteria as a bio-solar factory for photosynthetic production of a wide range of biofuels and commodity chemicals.

  20. Meganucleases Revolutionize the Production of Genetically Engineered Pigs for the Study of Human Diseases.

    Science.gov (United States)

    Redel, Bethany K; Prather, Randall S

    2016-04-01

    Animal models of human diseases are critically necessary for developing an in-depth knowledge of disease development and progression. In addition, animal models are vital to the development of potential treatments or even cures for human diseases. Pigs are exceptional models as their size, physiology, and genetics are closer to that of humans than rodents. In this review, we discuss the use of pigs in human translational research and the evolving technology that has increased the efficiency of genetically engineering pigs. With the emergence of the clustered, regularly interspaced, short palindromic repeat (CRISPR)/CRISPR-associated (Cas) protein 9 system technology, the cost and time it takes to genetically engineer pigs has markedly decreased. We will also discuss the use of another meganuclease, the transcription activator-like effector nucleases , to produce pigs with severe combined immunodeficiency by developing targeted modifications of the recombination activating gene 2 (RAG2).RAG2mutant pigs may become excellent animals to facilitate the development of xenotransplantation, regenerative medicine, and tumor biology. The use of pig biomedical models is vital for furthering the knowledge of, and for treating human, diseases.

  1. Genetic Engineering and Sustainable Crop Disease Management: Opportunities for Case-by-Case Decision-Making

    Directory of Open Access Journals (Sweden)

    Paul Vincelli

    2016-05-01

    Full Text Available Genetic engineering (GE offers an expanding array of strategies for enhancing disease resistance of crop plants in sustainable ways, including the potential for reduced pesticide usage. Certain GE applications involve transgenesis, in some cases creating a metabolic pathway novel to the GE crop. In other cases, only cisgenessis is employed. In yet other cases, engineered genetic changes can be so minimal as to be indistinguishable from natural mutations. Thus, GE crops vary substantially and should be evaluated for risks, benefits, and social considerations on a case-by-case basis. Deployment of GE traits should be with an eye towards long-term sustainability; several options are discussed. Selected risks and concerns of GE are also considered, along with genome editing, a technology that greatly expands the capacity of molecular biologists to make more precise and targeted genetic edits. While GE is merely a suite of tools to supplement other breeding techniques, if wisely used, certain GE tools and applications can contribute to sustainability goals.

  2. Field application of a genetically engineered microorganism for polycyclic aromatic hydrocarbon bioremediation process monitoring and control

    Energy Technology Data Exchange (ETDEWEB)

    Sayler, G.S.; Cox, C.D.; Ripp, S.; Nivens, D.E.; Werner, C.; Ahn, Y.; Matrubutham, U. [Univ. of Tennessee, Knoxville, TN (United States); Burlage, R. [Oak Ridge National Lab., TN (United States). Environmental Sciences Div.

    1998-11-01

    On October 30, 1996, the US Environmental Protection Agency (EPA) commenced the first test release of genetically engineered microorganisms (GEMs) for use in bioremediation. The specific objectives of the investigation were multifaceted and include (1) testing the hypothesis that a GEM can be successfully introduced and maintained in a bioremediation process, (2) testing the concept of using, at the field scale, reporter organisms for direct bioremediation process monitoring and control, and (3) acquiring data that can be used in risk assessment decision making and protocol development for future field release applications of GEMs. The genetically engineered strain under investigation is Pseudomonas fluorescens strain HK44 (King et al., 1990). The original P. fluorescens parent strain was isolated from polycyclic aromatic hydrocarbon (PAH) contaminated manufactured gas plant soil. Thus, this bacterium is able to biodegrade naphthalene (as well as other substituted naphthalenes and other PAHs) and is able to function as a living bioluminescent reporter for the presence of naphthalene contamination, its bioavailability, and the functional process of biodegradation. A unique component of this field investigation was the availability of an array of large subsurface soil lysimeters. This article describes the experience associated with the release of a genetically modified microorganism, the lysimeter facility and its associated instrumentation, as well as representative data collected during the first eighteen months of operation.

  3. Genetic engineering of crops: a ray of hope for enhanced food security.

    Science.gov (United States)

    Gill, Sarvajeet Singh; Gill, Ritu; Tuteja, Renu; Tuteja, Narendra

    2014-01-01

    Crop improvement has been a basic and essential chase since organized cultivation of crops began thousands of years ago. Abiotic stresses as a whole are regarded as the crucial factors restricting the plant species to reach their full genetic potential to deliver desired productivity. The changing global climatic conditions are making them worse and pointing toward food insecurity. Agriculture biotechnology or genetic engineering has allowed us to look into and understand the complex nature of abiotic stresses and measures to improve the crop productivity under adverse conditions. Various candidate genes have been identified and transformed in model plants as well as agriculturally important crop plants to develop abiotic stress-tolerant plants for crop improvement. The views presented here are an attempt toward realizing the potential of genetic engineering for improving crops to better tolerate abiotic stresses in the era of climate change, which is now essential for global food security. There is great urgency in speeding up crop improvement programs that can use modern biotechnological tools in addition to current breeding practices for providing enhanced food security.

  4. Genetically engineered K cells provide sufficient insulin to correct hyperglycemia in a nude murine model

    Institute of Scientific and Technical Information of China (English)

    Yiqun Zhang; Liqing Yao; Kuntang Shen; Meidong Xu; Pinghong Zhou; Weige Yang; Xinyuan Liu; Xinyu Qin

    2008-01-01

    A gene therapy-based treatment of type 1 diabetes mellitus requires the development of a surrogate β cell that can synthesize and secrete functionally active insulin in response to physiologically relevant changes in ambient glucose levels. In this study, the murine enteroendocrine cell line STC-1 was genetically modified by stable transfection. Two clone cells were selected (STC-1-2 and STC-1-14) that secreted the highest levels of insulin among the 22 clones expressing insulin from 0 to 157.2 μIU/ml/106 cells/d. After glucose concentration in the culture medium was increased from 1 mM to 10 mM, secreted insulin rose from 40.3±0.8 to 56.3±3.2 μIU/ml (STC-1-2), and from 10.8±0.8 to 23.6±2.3 μIU/ml (STC-1-14). After STC-1-14 cells were implanted into diabetic nude mice, their blood glucose levels were reduced to normal. Body weight loss was also ameliorated. Our data suggested that genetically engineered K cells secrete active insulin in a glucose-regulated manner, and in vivo study showed that hyperglycemia could be reversed by implantation of the cells, suggesting that the use of genetically engineered K cells to express human insulin might provide a glucose-regulated approach to treat diabetic hyperglycemia.

  5. Biomimetic self-templating optical structures fabricated by genetically engineered M13 bacteriophage.

    Science.gov (United States)

    Kim, Won-Geun; Song, Hyerin; Kim, Chuntae; Moon, Jong-Sik; Kim, Kyujung; Lee, Seung-Wuk; Oh, Jin-Woo

    2016-11-15

    Here, we describe a highly sensitive and selective surface plasmon resonance sensor system by utilizing self-assembly of genetically engineered M13 bacteriophage. About 2700 copies of genetically expressed peptide copies give superior selectivity and sensitivity to M13 phage-based SPR sensor. Furthermore, the sensitivity of the M13 phage-based SPR sensor was enhanced due to the aligning of receptor matrix in specific direction. Incorporation of specific binding peptide (His Pro Gln: HPQ) gives M13 bacteriophage high selectivity for the streptavidin. Our M13 phage-based SPR sensor takes advantage of simplicity of self-assembly compared with relatively complex photolithography techniques or chemical conjugations. Additionally, designed structure which is composed of functionalized M13 bacteriophage can simultaneously improve the sensitivity and selectivity of SPR sensor evidently. By taking advantages of the genetic engineering and self-assembly, we propose the simple method for fabricating novel M13 phage-based SPR sensor system which has a high sensitivity and high selectivity. Copyright © 2016 Elsevier B.V. All rights reserved.

  6. Improving itaconic acid production through genetic engineering of an industrial Aspergillus terreus strain.

    Science.gov (United States)

    Huang, Xuenian; Lu, Xuefeng; Li, Yueming; Li, Xia; Li, Jian-Jun

    2014-08-11

    Itaconic acid, which has been declared to be one of the most promising and flexible building blocks, is currently used as monomer or co-monomer in the polymer industry, and produced commercially by Aspergillus terreus. However, the production level of itaconic acid hasn't been improved in the past 40 years, and mutagenesis is still the main strategy to improve itaconate productivity. The genetic engineering approach hasn't been applied in industrial A. terreus strains to increase itaconic acid production. In this study, the genes closely related to itaconic acid production, including cadA, mfsA, mttA, ATEG_09969, gpdA, ATEG_01954, acoA, mt-pfkA and citA, were identified and overexpressed in an industrial A. terreus strain respectively. Overexpression of the genes cadA (cis-aconitate decarboxylase) and mfsA (Major Facilitator Superfamily Transporter) enhanced the itaconate production level by 9.4% and 5.1% in shake flasks respectively. Overexpression of other genes showed varied effects on itaconate production. The titers of other organic acids were affected by the introduced genes to different extent. Itaconic acid production could be improved through genetic engineering of the industrially used A. terreus strain. We have identified some important genes such as cadA and mfsA, whose overexpression led to the increased itaconate productivity, and successfully developed a strategy to establish a highly efficient microbial cell factory for itaconate protuction. Our results will provide a guide for further enhancement of the itaconic acid production level through genetic engineering in future.

  7. Construction and Expression of β-galactosidase Genetically Engineered Lactococcus lactis

    Institute of Scientific and Technical Information of China (English)

    吕晓英; 张朝武; 裴晓方; 刘祥; 余倩; 刘衡川

    2004-01-01

    Our objective is to solve the lactose malabsorption and intolerance of human beings by combining mlcro-ecology path with genetic engineering technique. Plasmid pMG36e was used to clone and express a β-galactosidase gene from L.delbrueckii bulgaricus strain 1. 1480 in the Lactococcus lactis subsp, cremoris MG1363 and Lactococcus lactis subsp. lactis IL1403. The recombinant plasmid was preserved and proliferated in Escherichia coli ( E. coli) JM109, and transformed into MG1363 and 1L1403 by electroporation. The protein expression was studied. (1) The bifidobacterium culture medium (BBL) was suitable for the growth of the strain 1. 1480. (2) With 13 amino acids at the N-terminus from the vector, β-galactosidase fusion protein (which retained the enzyme activity) could be successfully expressed in E. coli JM109, MG1363 and IL1403, but the expression quantity was larger in the former than in the latter two. (3) The SD sequence designed could be successfully recognized by both the E. coli and the Lactococcus lactis, but the expression level of the non-fusion β-galac-tosidase protein was lower than that of the fusion protein in the same host. The β-galactosidase genetically engineered E.coli JM109 is a useful tool to produce this enzyme in vitro. The signal peptide of the usp45 protein from the Lactococcus lactis can be added before the promoter sequence to promote β-galactosidase secretion from Lactococcus lactis. The potential application of the β-galactosidase genetically engineered MG1363 and IL1403 to cure the lactose malabsorption and lactose intolerance in both health food and medicine is promising。

  8. Construction and characterization of VL-VH tail-parallel genetically engineered antibodies against staphylococcal enterotoxins.

    Science.gov (United States)

    He, Xianzhi; Zhang, Lei; Liu, Pengchong; Liu, Li; Deng, Hui; Huang, Jinhai

    2015-03-01

    Staphylococcal enterotoxins (SEs) produced by Staphylococcus aureus have increasingly given rise to human health and food safety. Genetically engineered small molecular antibody is a useful tool in immuno-detection and treatment for clinical illness caused by SEs. In this study, we constructed the V(L)-V(H) tail-parallel genetically engineered antibody against SEs by using the repertoire of rearranged germ-line immunoglobulin variable region genes. Total RNA were extracted from six hybridoma cell lines that stably express anti-SEs antibodies. The variable region genes of light chain (V(L)) and heavy chain (V(H)) were cloned by reverse transcription PCR, and their classical murine antibody structure and functional V(D)J gene rearrangement were analyzed. To construct the eukaryotic V(H)-V(L) tail-parallel co-expression vectors based on the "5'-V(H)-ivs-IRES-V(L)-3'" mode, the ivs-IRES fragment and V(L) genes were spliced by two-step overlap extension PCR, and then, the recombined gene fragment and V(H) genes were inserted into the pcDNA3.1(+) expression vector sequentially. And then the constructed eukaryotic expression clones termed as p2C2HILO and p5C12HILO were transfected into baby hamster kidney 21 cell line, respectively. Two clonal cell lines stably expressing V(L)-V(H) tail-parallel antibodies against SEs were obtained, and the antibodies that expressed intracytoplasma were evaluated by enzyme-linked immunosorbent assay, immunofluorescence assay, and flow cytometry. SEs can stimulate the expression of some chemokines and chemokine receptors in porcine IPEC-J2 cells; mRNA transcription level of four chemokines and chemokine receptors can be blocked by the recombinant SE antibody prepared in this study. Our results showed that it is possible to get functional V(L)-V(H) tail-parallel genetically engineered antibodies in same vector using eukaryotic expression system.

  9. Optimal in silico target gene deletion through nonlinear programming for genetic engineering.

    Science.gov (United States)

    Hong, Chung-Chien; Song, Mingzhou

    2010-02-24

    Optimal selection of multiple regulatory genes, known as targets, for deletion to enhance or suppress the activities of downstream genes or metabolites is an important problem in genetic engineering. Such problems become more feasible to address in silico due to the availability of more realistic dynamical system models of gene regulatory and metabolic networks. The goal of the computational problem is to search for a subset of genes to knock out so that the activity of a downstream gene or a metabolite is optimized. Based on discrete dynamical system modeling of gene regulatory networks, an integer programming problem is formulated for the optimal in silico target gene deletion problem. In the first result, the integer programming problem is proved to be NP-hard and equivalent to a nonlinear programming problem. In the second result, a heuristic algorithm, called GKONP, is designed to approximate the optimal solution, involving an approach to prune insignificant terms in the objective function, and the parallel differential evolution algorithm. In the third result, the effectiveness of the GKONP algorithm is demonstrated by applying it to a discrete dynamical system model of the yeast pheromone pathways. The empirical accuracy and time efficiency are assessed in comparison to an optimal, but exhaustive search strategy. Although the in silico target gene deletion problem has enormous potential applications in genetic engineering, one must overcome the computational challenge due to its NP-hardness. The presented solution, which has been demonstrated to approximate the optimal solution in a practical amount of time, is among the few that address the computational challenge. In the experiment on the yeast pheromone pathways, the identified best subset of genes for deletion showed advantage over genes that were selected empirically. Once validated in vivo, the optimal target genes are expected to achieve higher genetic engineering effectiveness than a trial

  10. Optimal in silico target gene deletion through nonlinear programming for genetic engineering.

    Directory of Open Access Journals (Sweden)

    Chung-Chien Hong

    Full Text Available BACKGROUND: Optimal selection of multiple regulatory genes, known as targets, for deletion to enhance or suppress the activities of downstream genes or metabolites is an important problem in genetic engineering. Such problems become more feasible to address in silico due to the availability of more realistic dynamical system models of gene regulatory and metabolic networks. The goal of the computational problem is to search for a subset of genes to knock out so that the activity of a downstream gene or a metabolite is optimized. METHODOLOGY/PRINCIPAL FINDINGS: Based on discrete dynamical system modeling of gene regulatory networks, an integer programming problem is formulated for the optimal in silico target gene deletion problem. In the first result, the integer programming problem is proved to be NP-hard and equivalent to a nonlinear programming problem. In the second result, a heuristic algorithm, called GKONP, is designed to approximate the optimal solution, involving an approach to prune insignificant terms in the objective function, and the parallel differential evolution algorithm. In the third result, the effectiveness of the GKONP algorithm is demonstrated by applying it to a discrete dynamical system model of the yeast pheromone pathways. The empirical accuracy and time efficiency are assessed in comparison to an optimal, but exhaustive search strategy. SIGNIFICANCE: Although the in silico target gene deletion problem has enormous potential applications in genetic engineering, one must overcome the computational challenge due to its NP-hardness. The presented solution, which has been demonstrated to approximate the optimal solution in a practical amount of time, is among the few that address the computational challenge. In the experiment on the yeast pheromone pathways, the identified best subset of genes for deletion showed advantage over genes that were selected empirically. Once validated in vivo, the optimal target genes are

  11. Field cage studies and progressive evaluation of genetically-engineered mosquitoes.

    Directory of Open Access Journals (Sweden)

    Luca Facchinelli

    Full Text Available A genetically-engineered strain of the dengue mosquito vector Aedes aegypti, designated OX3604C, was evaluated in large outdoor cage trials for its potential to improve dengue prevention efforts by inducing population suppression. OX3604C is engineered with a repressible genetic construct that causes a female-specific flightless phenotype. Wild-type females that mate with homozygous OX3604C males will not produce reproductive female offspring. Weekly introductions of OX3604C males eliminated all three targeted Ae. aegypti populations after 10-20 weeks in a previous laboratory cage experiment. As part of the phased, progressive evaluation of this technology, we carried out an assessment in large outdoor field enclosures in dengue endemic southern Mexico.OX3604C males were introduced weekly into field cages containing stable target populations, initially at 10:1 ratios. Statistically significant target population decreases were detected in 4 of 5 treatment cages after 17 weeks, but none of the treatment populations were eliminated. Mating competitiveness experiments, carried out to explore the discrepancy between lab and field cage results revealed a maximum mating disadvantage of up 59.1% for OX3604C males, which accounted for a significant part of the 97% fitness cost predicted by a mathematical model to be necessary to produce the field cage results.Our results indicate that OX3604C may not be effective in large-scale releases. A strain with the same transgene that is not encumbered by a large mating disadvantage, however, could have improved prospects for dengue prevention. Insights from large outdoor cage experiments may provide an important part of the progressive, stepwise evaluation of genetically-engineered mosquitoes.

  12. Porcine Fetal Bone Marrow Mesenchymal Stem Cells in Cartilage Tissue Engineering%胎猪BMSC体外构建软骨的实验研究

    Institute of Scientific and Technical Information of China (English)

    刘李娜; 何爱娟; 周广东; 曹卫刚

    2013-01-01

    Objective To investigate the optimal seed cell for cartilage engineering by comparing the chondrogenesis capability of porcine fetal bone marrow mesenchymal stem cells (fBMSCs) and adult porcine BMSCs (aBMSCs). Methods Fetal pigs with gestational age of 70 days were obtained by the uterine-incision delivery, and primary fBMSCs were isolated from the bone marrow. Primary aBMSCs were isolated from the bone marrow which was aspirated from iliac. Cell morphology of the two kinds of cells at passage 3 were observed after in vitro proliferation. The two kinds of cells at passages 3 were characterized by their osteogenic, adipogenic and chondrogenic differentiation potential. Then the fBMSCs and aBMSCs were separately seeded onto a polyglycolic acid/polylactic acid (PGA/PLA) scaffold with the concentration of 1 ×108 cells/mL. All specimens were harvested after 8 weeks' culture in vitro. Gross observation, glycosaminoglycan (GAG) quantification, total collagen quantification and histology were used to compare related characteristic differences of engineered cartilage formed by the two kinds of cells. Results fBMSCs had better proliferation and multiple differentiation capacity than aBMSCs. The two kinds of cells both formed mature cartilage after 8 weeks of culture in vitro, and the engineered cartilage of aBMSCs group had better appearance. The GAG content and total collagen content of the cartilage formed by fBMSCs were both higher than the cartilage formed by aBMSCs (P<0.01). Histology and immunohistochemistry demonstrated that the cartilage formed by fBMSCs have more compact tissue structure. The cartilage matrix staining of cartilage formed by fBMSCs were stronger than that of cartilage formed by aBMSCs. Conclusion The fBMSCs seems to be the optimal seed cells for cartilage tissue engineering.%目的比较胎猪骨髓间充质干细胞(Bone Marrow Mesenchymal Stem Cells,BMSCs)和成年猪BMSCs构建软骨能力的差异,寻找合适的同种异体组织工

  13. Optimization of a Reduced Chemical Kinetic Model for HCCI Engine Simulations by Micro-Genetic Algorithm

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    A reduced chemical kinetic model (44 species and 72 reactions) for the homogeneous charge compression ignition (HCCI) combustion of n-heptane was optimized to improve its autoignition predictions under different engine operating conditions. The seven kinetic parameters of the optimized model were determined by using the combination of a micro-genetic algorithm optimization methodology and the SENKIN program of CHEMKIN chemical kinetics software package. The optimization was performed within the range of equivalence ratios 0.2-1.2, initial temperature 310-375 K and initial pressure 0.1-0.3 MPa. The engine simulations show that the optimized model agrees better with the detailed chemical kinetic model (544 species and 2 446 reactions) than the original model does.

  14. Conflicts of interest among committee members in the National Academies’ genetically engineered crop study

    Science.gov (United States)

    2017-01-01

    The National Academies of Sciences, Engineering and Medicine (NASEM) publishes numerous reports each year that are received with high esteem by the scientific community and public policy makers. The NASEM has internal standards for selecting committee members that author its reports, mostly from academia, and vetting conflicts of interest. This study examines whether there were any financial conflicts of interest (COIs) among the twenty invited committee members who wrote the 2016 report on genetically engineered (GE) crops. Our results showed that six panel members had one or more reportable financial COIs, none of which were disclosed in the report. We also report on institutional COIs held by the NASEM related to the report. The difference between our findings and the NASEM reporting standards are discussed. PMID:28245228

  15. Engineering antigen-specific T cells from genetically modified human hematopoietic stem cells in immunodeficient mice.

    Directory of Open Access Journals (Sweden)

    Scott G Kitchen

    Full Text Available There is a desperate need for effective therapies to fight chronic viral infections. The immune response is normally fastidious at controlling the majority of viral infections and a therapeutic strategy aimed at reestablishing immune control represents a potentially powerful approach towards treating persistent viral infections. We examined the potential of genetically programming human hematopoietic stem cells to generate mature CD8+ cytotoxic T lymphocytes that express a molecularly cloned, "transgenic" human anti-HIV T cell receptor (TCR. Anti-HIV TCR transduction of human hematopoietic stem cells directed the maturation of a large population of polyfunctional, HIV-specific CD8+ cells capable of recognizing and killing viral antigen-presenting cells. Thus, through this proof-of-concept we propose that genetic engineering of human hematopoietic stem cells will allow the tailoring of effector T cell responses to fight HIV infection or other diseases that are characterized by the loss of immune control.

  16. Genetic engineering of baker's and wine yeasts using formaldehyde hyperresistance-mediating plasmids

    Directory of Open Access Journals (Sweden)

    Schmidt M.

    1997-01-01

    Full Text Available Yeast multi-copy vectors carrying the formaldehyde-resistance marker gene SFA have proved to be a valuable tool for research on industrially used strains of Saccharomyces cerevisiae. The genetics of these strains is often poorly understood, and for various reasons it is not possible to simply subject these strains to protocols of genetic engineering that have been established for laboratory strains of S. cerevisiae. We tested our vectors and protocols using 10 randomly picked baker's and wine yeasts all of which could be transformed by a simple protocol with vectors conferring hyperresistance to formaldehyde. The application of formaldehyde as a selecting agent also offers the advantage of its biodegradation to CO2 during fermentation, i.e., the selecting agent will be consumed and therefore its removal during down-stream processing is not necessary. Thus, this vector provides an expression system which is simple to apply and inexpensive to use

  17. Phytosequestration: Carbon biosequestration by plants and the prospects of genetic engineering

    Energy Technology Data Exchange (ETDEWEB)

    Jansson, C.; Wullschleger, S.D.; Kalluri, U.C.; Tuskan, G.A.

    2010-07-15

    Photosynthetic assimilation of atmospheric carbon dioxide by land plants offers the underpinnings for terrestrial carbon (C) sequestration. A proportion of the C captured in plant biomass is partitioned to roots, where it enters the pools of soil organic C and soil inorganic C and can be sequestered for millennia. Bioenergy crops serve the dual role of providing biofuel that offsets fossil-fuel greenhouse gas (GHG) emissions and sequestering C in the soil through extensive root systems. Carbon captured in plant biomass can also contribute to C sequestration through the deliberate addition of biochar to soil, wood burial, or the use of durable plant products. Increasing our understanding of plant, microbial, and soil biology, and harnessing the benefits of traditional genetics and genetic engineering, will help us fully realize the GHG mitigation potential of phytosequestration.

  18. The genetics of murine Hox loci: TAMERE, STRING, and PANTHERE to engineer chromosome variants.

    Science.gov (United States)

    Tschopp, Patrick; Duboule, Denis

    2014-01-01

    Following their duplications at the base of the vertebrate clade, Hox gene clusters underwent remarkable sub- and neo-functionalization events. Many of these evolutionary innovations can be associated with changes in the transcriptional regulation of their genes, where an intricate relationship between the structure of the gene cluster and the architecture of the surrounding genomic landscape is at play. Here, we report on a portfolio of in vivo genome engineering strategies in mice, which have been used to probe and decipher the genetic and molecular underpinnings of the complex regulatory mechanisms implemented at these loci.

  19. Illuminating cancer systems with genetically engineered mouse models and coupled luciferase reporters in vivo.

    Science.gov (United States)

    Kocher, Brandon; Piwnica-Worms, David

    2013-06-01

    Bioluminescent imaging (BLI) is a powerful noninvasive tool that has dramatically accelerated the in vivo interrogation of cancer systems and longitudinal analysis of mouse models of cancer over the past decade. Various luciferase enzymes have been genetically engineered into mouse models (GEMM) of cancer, which permit investigation of cellular and molecular events associated with oncogenic transcription, posttranslational processing, protein-protein interactions, transformation, and oncogene addiction in live cells and animals. Luciferase-coupled GEMMs ultimately serve as a noninvasive, repetitive, longitudinal, and physiologic means by which cancer systems and therapeutic responses can be investigated accurately within the autochthonous context of a living animal.

  20. Osmoregulation Mechanism of Drought Stress and Genetic Engineering Strategies for Improving Drought Resistance in Plants

    Institute of Scientific and Technical Information of China (English)

    Du Jinyou; Chen Xiaoyang; Li Wei; Gao Qiong

    2004-01-01

    Drought, one of the main adverse environmental factors, obviously affected plant growth and development. Many adaptive strategies have been developed in plants for coping with drought or water stress, among which osmoregulation is one of the important factors of plant drought tolerance. Many substances play important roles in plant osmoregulation for drought resistance, including proline, glycine betaine, Lea proteins and soluble sugars such as levan, trehalose, sucrose, etc. The osmoregulation mechanism and the genetic engineering of plant drought-tolerance are reviewed in this paper.

  1. Crystals of Serum Albumin for Use in Genetic Engineering and Rational Drug Design

    Science.gov (United States)

    Carter, Daniel C. (Inventor)

    1996-01-01

    Serum albumin crystal forms have been produced which exhibit superior x-ray diffraction quality. The crystals are produced from both recombinant and wild-type human serum albumin, canine, and baboon serum albumin and allow the performance of drug-binding studies as well as genetic engineering studies. The crystals are grown from solutions of polyethylene glycol or ammonium sulphate within prescribed limits during growth times from one to several weeks and include the following space groups: P2(sub 1), C2, P1.

  2. Illuminating p53 function in cancer with genetically engineered mouse models

    OpenAIRE

    2014-01-01

    The key role of the p53 protein in tumor suppression is highlighted by its frequent mutation in human cancers and by the completely penetrant cancer predisposition of p53 null mice. Beyond providing definitive evidence for the critical function of p53 in tumor suppression, genetically engineered mouse models have offered numerous additional insights into p53 function. p53 knock-in mice expressing tumor-derived p53 mutants have revealed that these mutants display gain-of-function activities th...

  3. Can genetic engineering of lignin deposition be accomplished without an unacceptable yield penalty?

    Science.gov (United States)

    Bonawitz, Nicholas D; Chapple, Clint

    2013-04-01

    The secondary cell wall polymer lignin impedes the extraction of fermentable sugars from biomass, and has been one of the major impediments in the development of cost-effective biofuel technologies. Unfortunately, attempts to genetically engineer lignin biosynthesis frequently result in dwarfing or developmental abnormalities of unknown cause, thus limiting the benefits of increased fermentable sugar yield. In this brief review, we explore some of the possible mechanisms that could underlie this poorly understood phenomenon, with the expectation that an understanding of the cause of dwarfing in lignin biosynthetic mutants and transgenic plants could lead to new strategies for the development of improved bioenergy feedstocks. Copyright © 2012 Elsevier Ltd. All rights reserved.

  4. Analysis and design of a genetic circuit for dynamic metabolic engineering.

    Science.gov (United States)

    Anesiadis, Nikolaos; Kobayashi, Hideki; Cluett, William R; Mahadevan, Radhakrishnan

    2013-08-16

    Recent advances in synthetic biology have equipped us with new tools for bioprocess optimization at the genetic level. Previously, we have presented an integrated in silico design for the dynamic control of gene expression based on a density-sensing unit and a genetic toggle switch. In the present paper, analysis of a serine-producing Escherichia coli mutant shows that an instantaneous ON-OFF switch leads to a maximum theoretical productivity improvement of 29.6% compared to the mutant. To further the design, global sensitivity analysis is applied here to a mathematical model of serine production in E. coli coupled with a genetic circuit. The model of the quorum sensing and the toggle switch involves 13 parameters of which 3 are identified as having a significant effect on serine concentration. Simulations conducted in this reduced parameter space further identified the optimal ranges for these 3 key parameters to achieve productivity values close to the maximum theoretical values. This analysis can now be used to guide the experimental implementation of a dynamic metabolic engineering strategy and reduce the time required to design the genetic circuit components.

  5. Genetic variation changes the interactions between the parasitic plant-ecosystem engineer Rhinanthus and its hosts.

    Science.gov (United States)

    Rowntree, Jennifer K; Cameron, Duncan D; Preziosi, Richard F

    2011-05-12

    Within-species genetic variation is a potent factor influencing between-species interactions and community-level structure. Species of the hemi-parasitic plant genus Rhinanthus act as ecosystem engineers, significantly altering above- and below-ground community structure in grasslands. Here, we show the importance of genotypic variation within a single host species (barley-Hordeum vulgare), and population-level variation among two species of parasite (Rhinanthus minor and Rhinanthus angustifolius) on the outcome of parasite infection for both partners. We measured host fitness (number of seeds) and calculated parasite virulence as the difference in seed set between infected and uninfected hosts (the inverse of host tolerance). Virulence was determined by genetic variation within the host species and among the parasite species, but R. angustifolius was consistently more virulent than R. minor. The most tolerant host had the lowest inherent fitness and did not gain a fitness advantage over other infected hosts. We measured parasite size as a proxy for transmission ability (ability to infect further hosts) and host resistance. Parasite size depended on the specific combination of host genotype, parasite species and parasite population, and no species was consistently larger. We demonstrate that the outcome of infection by Rhinanthus depends not only on the host species, but also on the underlying genetics of both host and parasite. Thus, genetic variations within host and parasite are probably essential components of the ecosystem-altering effects of Rhinanthus.

  6. Design and testing of a synthetic biology framework for genetic engineering of Corynebacterium glutamicum

    Directory of Open Access Journals (Sweden)

    Ravasi Pablo

    2012-11-01

    Full Text Available Abstract Background Synthetic biology approaches can make a significant contribution to the advance of metabolic engineering by reducing the development time of recombinant organisms. However, most of synthetic biology tools have been developed for Escherichia coli. Here we provide a platform for rapid engineering of C. glutamicum, a microorganism of great industrial interest. This bacteria, used for decades for the fermentative production of amino acids, has recently been developed as a host for the production of several economically important compounds including metabolites and recombinant proteins because of its higher capacity of secretion compared to traditional bacterial hosts like E. coli. Thus, the development of modern molecular platforms may significantly contribute to establish C. glutamicum as a robust and versatile microbial factory. Results A plasmid based platform named pTGR was created where all the genetic components are flanked by unique restriction sites to both facilitate the evaluation of regulatory sequences and the assembly of constructs for the expression of multiple genes. The approach was validated by using reporter genes to test promoters, ribosome binding sites, and for the assembly of dual gene operons and gene clusters containing two transcriptional units. Combinatorial assembly of promoter (tac, cspB and sod and RBS (lacZ, cspB and sod elements with different strengths conferred clear differential gene expression of two reporter genes, eGFP and mCherry, thus allowing transcriptional “fine-tuning”of multiple genes. In addition, the platform allowed the rapid assembly of operons and genes clusters for co-expression of heterologous genes, a feature that may assist metabolic pathway engineering. Conclusions We anticipate that the pTGR platform will contribute to explore the potential of novel parts to regulate gene expression, and to facilitate the assembly of genetic circuits for metabolic engineering of C

  7. Is genetic engineering ever going to take off in forage, turf and bioenergy crop breeding?

    Science.gov (United States)

    Wang, Zeng-Yu; Brummer, E Charles

    2012-11-01

    Genetic engineering offers the opportunity to generate unique genetic variation that is either absent in the sexually compatible gene pool or has very low heritability. The generation of transgenic plants, coupled with breeding, has led to the production of widely used transgenic cultivars in several major cash crops, such as maize, soybean, cotton and canola. The process for regulatory approval of genetically engineered crops is slow and subject to extensive political interference. The situation in forage grasses and legumes is more complicated. Most widely grown forage, turf and bioenergy species (e.g. tall fescue, perennial ryegrass, switchgrass, alfalfa, white clover) are highly self-incompatible and outcrossing. Compared with inbreeding species, they have a high potential to pass their genes to adjacent plants. A major biosafety concern in these species is pollen-mediated transgene flow. Because human consumption is indirect, risk assessment of transgenic forage, turf and bioenergy species has focused on their environmental or ecological impacts. Although significant progress has been made in genetic modification of these species, commercialization of transgenic cultivars is very limited because of the stringent and costly regulatory requirements. To date, the only transgenic forage crop deregulated in the US is 'Roundup Ready' (RR) alfalfa. The approval process for RR alfalfa was complicated, involving several rounds of regulation, deregulation and re-regulation. Nevertheless, commercialization of RR alfalfa is an important step forward in regulatory approval of a perennial outcrossing forage crop. As additional transgenic forage, turf and bioenergy crops are generated and tested, different strategies have been developed to meet regulatory requirements. Recent progress in risk assessment and deregulation of transgenic forage and turf species is summarized and discussed.

  8. Pancreatic mesenchyme regulates epithelial organogenesis throughout development.

    Directory of Open Access Journals (Sweden)

    Limor Landsman

    2011-09-01

    Full Text Available The developing pancreatic epithelium gives rise to all endocrine and exocrine cells of the mature organ. During organogenesis, the epithelial cells receive essential signals from the overlying mesenchyme. Previous studies, focusing on ex vivo tissue explants or complete knockout mice, have identified an important role for the mesenchyme in regulating the expansion of progenitor cells in the early pancreas epithelium. However, due to the lack of genetic tools directing expression specifically to the mesenchyme, the potential roles of this supporting tissue in vivo, especially in guiding later stages of pancreas organogenesis, have not been elucidated. We employed transgenic tools and fetal surgical techniques to ablate mesenchyme via Cre-mediated mesenchymal expression of Diphtheria Toxin (DT at the onset of pancreas formation, and at later developmental stages via in utero injection of DT into transgenic mice expressing the Diphtheria Toxin receptor (DTR in this tissue. Our results demonstrate that mesenchymal cells regulate pancreatic growth and branching at both early and late developmental stages by supporting proliferation of precursors and differentiated cells, respectively. Interestingly, while cell differentiation was not affected, the expansion of both the endocrine and exocrine compartments was equally impaired. To further elucidate signals required for mesenchymal cell function, we eliminated β-catenin signaling and determined that it is a critical pathway in regulating mesenchyme survival and growth. Our study presents the first in vivo evidence that the embryonic mesenchyme provides critical signals to the epithelium throughout pancreas organogenesis. The findings are novel and relevant as they indicate a critical role for the mesenchyme during late expansion of endocrine and exocrine compartments. In addition, our results provide a molecular mechanism for mesenchymal expansion and survival by identifying β-catenin signaling as an

  9. Identification of stable reference genes for gene expression analysis of three-dimensional cultivated human bone marrow-derived mesenchymal stromal cells for bone tissue engineering.

    Science.gov (United States)

    Rauh, Juliane; Jacobi, Angela; Stiehler, Maik

    2015-02-01

    The principles of tissue engineering (TE) are widely used for bone regeneration concepts. Three-dimensional (3D) cultivation of autologous human mesenchymal stromal cells (MSCs) on porous scaffolds is the basic prerequisite to generate newly formed bone tissue. Quantitative reverse transcription polymerase chain reaction (qRT-PCR) is a specific and sensitive analytical tool for the measurement of mRNA-levels in cells or tissues. For an accurate quantification of gene expression levels, stably expressed reference genes (RGs) are essential to obtain reliable results. Since the 3D environment can affect a cell's morphology, proliferation, and gene expression profile compared with two-dimensional (2D) cultivation, there is a need to identify robust RGs for the quantification of gene expression. So far, this issue has not been adequately investigated. The aim of this study was to identify the most stably expressed RGs for gene expression analysis of 3D-cultivated human bone marrow-derived MSCs (BM-MSCs). For this, we analyzed the gene expression levels of n=31 RGs in 3D-cultivated human BM-MSCs from six different donors compared with conventional 2D cultivation using qRT-PCR. MSCs isolated from bone marrow aspirates were cultivated on human cancellous bone cube scaffolds for 14 days. Osteogenic differentiation was assessed by cell-specific alkaline phosphatase (ALP) activity and expression of osteogenic marker genes. Expression levels of potential reference and target genes were quantified using commercially available TaqMan(®) assays. mRNA expression stability of RGs was determined by calculating the coefficient of variation (CV) and using the algorithms of geNorm and NormFinder. Using both algorithms, we identified TATA box binding protein (TBP), transferrin receptor (p90, CD71) (TFRC), and hypoxanthine phosphoribosyltransferase 1 (HPRT1) as the most stably expressed RGs in 3D-cultivated BM-MSCs. Notably, genes that are routinely used as RGs, for example, beta actin

  10. Evaluation of terrestrial microcosms for detection, fate, and survival analysis of genetically engineered microorganisms and their recombinant genetic material

    Energy Technology Data Exchange (ETDEWEB)

    Fredrickson, J.K.; Seidler, R.J.

    1989-02-01

    The research included in this document represents the current scientific information available regarding the applicability of terrestrial microcosms and related methodologies for evaluating detection methods and the fate and survival of microorganisms in the environment. The three terrestrial microcosms described in this document were used to evaluate the survival and fate of recombinant bacteria in soils and in association with plant surfaces and insects and their transport through soil with percolating water and root systems, and to test new methods and procedures to improve detection and enumeration of bacteria in soil. Simple (potting soil composed of peat mix and perlite, lacking environmental control and monitoring) and complex microcosms (agricultural soil with partial control and monitoring of environmental conditions) were demonstrated to be useful tools for preliminary assessments of microbial viability in terrestrial ecosystems. These studies evaluated the survival patterns of Enterobacter cloacae (pBR322) in soil and on plant surfaces and the ingestion of this same microorganism by cutworms and survival in the foregut and frass. The Versacore microcosm design was used to monitor the fate and competitiveness of genetically engineered bacteria in soil. Both selective media and gene probes were used successfully to follow the fate of two recombinant Pseudomonas sp. introduced into Versacore microcosms. Intact soil-core microcosms were employed to evaluate the fate and transport of genetically altered Azospirillum sp. and Pseudomonas sp. in soil and the plant rhizosphere. The usefulness of these various microcosms as a tool for risk assessment is underscored by the ease in obtaining soil from a proposed field release site to evaluate subsequent GEM fate and survival.

  11. Progress in genetic engineering of peanut (Arachis hypogaea L.)--a review.

    Science.gov (United States)

    Krishna, Gaurav; Singh, Birendra K; Kim, Eun-Ki; Morya, Vivek K; Ramteke, Pramod W

    2015-02-01

    Peanut (Arachis hypogaea L.) is a major species of the family, Leguminosae, and economically important not only for vegetable oil but as a source of proteins, minerals and vitamins. It is widely grown in the semi-arid tropics and plays a role in the world agricultural economy. Peanut production and productivity is constrained by several biotic (insect pests and diseases) and abiotic (drought, salinity, water logging and temperature aberrations) stresses, as a result of which crop experiences serious economic losses. Genetic engineering techniques such as Agrobacterium tumefaciens and DNA-bombardment-mediated transformation are used as powerful tools to complement conventional breeding and expedite peanut improvement by the introduction of agronomically useful traits in high-yield background. Resistance to several fungal, virus and insect pest have been achieved through variety of approaches ranging from gene coding for cell wall component, pathogenesis-related proteins, oxalate oxidase, bacterial chloroperoxidase, coat proteins, RNA interference, crystal proteins etc. To develop transgenic plants withstanding major abiotic stresses, genes coding transcription factors for drought and salinity, cytokinin biosynthesis, nucleic acid processing, ion antiporter and human antiapoptotic have been used. Moreover, peanut has also been used in vaccine production for the control of several animal diseases. In addition to above, this study also presents a comprehensive account on the influence of some important factors on peanut genetic engineering. Future research thrusts not only suggest the use of different approaches for higher expression of transgene(s) but also provide a way forward for the improvement of crops.

  12. Genetically engineered alginate lyase-PEG conjugates exhibit enhanced catalytic function and reduced immunoreactivity.

    Directory of Open Access Journals (Sweden)

    John W Lamppa

    Full Text Available Alginate lyase enzymes represent prospective biotherapeutic agents for treating bacterial infections, particularly in the cystic fibrosis airway. To effectively deimmunize one therapeutic candidate while maintaining high level catalytic proficiency, a combined genetic engineering-PEGylation strategy was implemented. Rationally designed, site-specific PEGylation variants were constructed by orthogonal maleimide-thiol coupling chemistry. In contrast to random PEGylation of the enzyme by NHS-ester mediated chemistry, controlled mono-PEGylation of A1-III alginate lyase produced a conjugate that maintained wild type levels of activity towards a model substrate. Significantly, the PEGylated variant exhibited enhanced solution phase kinetics with bacterial alginate, the ultimate therapeutic target. The immunoreactivity of the PEGylated enzyme was compared to a wild type control using in vitro binding studies with both enzyme-specific antibodies, from immunized New Zealand white rabbits, and a single chain antibody library, derived from a human volunteer. In both cases, the PEGylated enzyme was found to be substantially less immunoreactive. Underscoring the enzyme's potential for practical utility, >90% of adherent, mucoid, Pseudomonas aeruginosa biofilms were removed from abiotic surfaces following a one hour treatment with the PEGylated variant, whereas the wild type enzyme removed only 75% of biofilms in parallel studies. In aggregate, these results demonstrate that site-specific mono-PEGylation of genetically engineered A1-III alginate lyase yielded an enzyme with enhanced performance relative to therapeutically relevant metrics.

  13. Genetic engineering: a promising tool to engender physiological, biochemical and molecular stress resilience in green microalgae

    Directory of Open Access Journals (Sweden)

    Freddy eGuiheneuf

    2016-03-01

    Full Text Available As we march into the 21st century, the prevailing scenario of depleting energy resources, global warming and ever increasing issues of human health and food security will quadruple. In this context, genetic and metabolic engineering of green microalgae complete the quest towards a continuum of environmentally clean fuel and food production. Evolutionarily related, but unlike land plants, microalgae need nominal land or water, and are best described as unicellular autotrophs using light energy to fix atmospheric CO2 into algal biomass, mitigating fossil CO2 pollution in the process. Remarkably, a feature innate to most microalgae is synthesis and accumulation of lipids (60–65% of dry weight, carbohydrates and secondary metabolites like pigments and vitamins, especially when grown under abiotic stress conditions. Particularly fruitful, such an application of abiotic stress factors like nitrogen starvation , salinity, heat shock etc. can be used in a biorefinery concept for production of multiple valuable products. The focus of this mini-review underlies metabolic reorientation practices and tolerance mechanisms as applied to green microalgae under specific stress stimuli for a sustainable pollution-free future. Moreover, we entail current progress on genetic engineering as a promising tool to grasp adaptive processes for improving strains with potential biotechnological interests.

  14. Enhanced atrazine removal using membrane bioreactor bioaugmented with genetically engineered microorganism

    Institute of Scientific and Technical Information of China (English)

    Chun LIU; Xia HUANG

    2008-01-01

    Bioaugmentation with genetically engineered microorganisms (GEMs) in a membrane bioreactor (MBR) for enhanced removal of recalcitrant pollutants was explored. An atrazine-degrading genetically engi-neered microorganism (GEM) with green fluorescent pro-tein was inoculated into an MBR and the effects of such a bioaugmentation strategy on atrazine removal were inves-tigated. The results show that atrazine removal was improved greatly in the bioaugmented MBR compared with a control system. After a start-up period of 6 days, average 94.7% of atrazine was removed in bioaugmented MBR when atrazine concentration of influent was 14.5 mg/L. The volu-metric removal rates increased linearly followed by atrazine loading increase and the maximum was 65.5 mg/(L·d). No negative effects were found on COD removal although carbon oxidation activity of bioaugmented sludge was lower than that of common sludge. After inoculation, adsorption to sludge flocs was favorable for GEM sur-vival. The GEM population size initially decreased shortly and then was kept constant at about 104-105 CFU/mL. Predation of micro-organisms played an important role in the decay of the GEM population. GEM leakage from MBR was less than 102 CFU/mL initially and was then undetectable. In contrast, in a conventionally activated sludge bioreactor (CAS), sludge bulking occurred possibly due to atrazine exposure, resulting in bioaugmentation failure and serious GEM leakage. So MBR was superior to CAS in atrazine bioaugmentation treatment using GEM.

  15. Recent advances to improve fermentative butanol production: genetic engineering and fermentation technology.

    Science.gov (United States)

    Zheng, Jin; Tashiro, Yukihiro; Wang, Qunhui; Sonomoto, Kenji

    2015-01-01

    Butanol has recently attracted attention as an alternative biofuel because of its various advantages over other biofuels. Many researchers have focused on butanol fermentation with renewable and sustainable resources, especially lignocellulosic materials, which has provided significant progress in butanol fermentation. However, there are still some drawbacks in butanol fermentation in terms of low butanol concentration and productivity, high cost of feedstock and product inhibition, which makes butanol fermentation less competitive than the production of other biofuels. These hurdles are being resolved in several ways. Genetic engineering is now available for improving butanol yield and butanol ratio through overexpression, knock out/down, and insertion of genes encoding key enzymes in the metabolic pathway of butanol fermentation. In addition, there are also many strategies to improve fermentation technology, such as multi-stage continuous fermentation, continuous fermentation integrated with immobilization and cell recycling, and the inclusion of additional organic acids or electron carriers to change metabolic flux. This review focuses on the most recent advances in butanol fermentation especially from the perspectives of genetic engineering and fermentation technology.

  16. Crop Resources Ethic in Plant Genetic Engineering and Fortune Transfer Between Generations

    Institute of Scientific and Technical Information of China (English)

    WANG Xiaowei; DING Guangzhou; LIANG Xueqing

    2006-01-01

    The relation between human and crop resources belongs to the ethic of resources exploitation. The purposes of discussing the ethic of crop resources are to protect the ecology and safety of crops, to gain sustainable development, furthermore, to choose and form the production structure that is favorable to saving crop resources and protecting the ecology of crops. Plant genetic engineering is the technology of molecule breeding of rearrangement of inheritance materials at the level of molecule directionally, of improving plant properties and of breeding high quality and yield varieties of crops. The prominent effects of the technology on the crop ecological system are human subjective factors increasing as well as violating the nature and intensifying the conflict between human being and nature.Therefore, in plant genetic engineering, crop resources exploitation should follow certain ethic principles. Under the theory of ethics of natural resources, by the means of biologioal statistics, the author systematically analyzed the possible model of crop resources transfer between generations as well as the transfer mode of magnitude of real materials and magnitude of value.

  17. Deconjugation of Bile Acids with Immobilized Genetically Engineered Lactobacillus plantarum 80(pCBH1

    Directory of Open Access Journals (Sweden)

    M. L. Jones

    2005-01-01

    Full Text Available Bile acids are important to normal human physiology. However, bile acids can be toxic when produced in pathologically high concentrations in hepatobileary and other diseases. This study shows that immobilized genetically engineered Lactobacillus plantarum 80 (pCBH1 (LP80 (pCBH1 can efficiently hydrolyze bile acids and establishes a basis for their use. Results show that immobilized LP80 (pCBH1 is able to effectively break down the conjugated bile acids into glycodeoxycholic acid (GDCA and taurodeoxycholic acid (TDCA with bile salt hydrolase (BSH activities of 0.17 and 0.07 μmol DCA/mg CDW/h, respectively. The deconjugation product, deoxycholic acid (DCA, was diminished by LP80 (pCBH1 within 4 h of initial BSH activity. This in-vitro study suggests that immobilized genetically engineered bacterial cells have important potential for deconjugation of bile acids for lowering of high levels of bile acids for therapy.

  18. Designing RNA-based genetic control systems for efficient production from engineered metabolic pathways.

    Science.gov (United States)

    Stevens, Jason T; Carothers, James M

    2015-02-20

    Engineered metabolic pathways can be augmented with dynamic regulatory controllers to increase production titers by minimizing toxicity and helping cells maintain homeostasis. We investigated the potential for dynamic RNA-based genetic control systems to increase production through simulation analysis of an engineered p-aminostyrene (p-AS) pathway in E. coli. To map the entire design space, we formulated 729 unique mechanistic models corresponding to all of the possible control topologies and mechanistic implementations in the system under study. Two thousand sampled simulations were performed for each of the 729 system designs to relate the potential effects of dynamic control to increases in p-AS production (total of 3 × 10(6) simulations). Our analysis indicates that dynamic control strategies employing aptazyme-regulated expression devices (aREDs) can yield >10-fold improvements over static control. We uncovered generalizable trends in successful control architectures and found that highly performing RNA-based control systems are experimentally tractable. Analyzing the metabolic control state space to predict optimal genetic control strategies promises to enhance the design of metabolic pathways.

  19. Stakeholder views on the creation and use of genetically-engineered animals in research.

    Science.gov (United States)

    Ormandy, Elisabeth H

    2016-05-01

    This interview-based study examined the diversity of views relating to the creation and use of genetically-engineered (GE) animals in biomedical science. Twenty Canadian participants (eight researchers, five research technicians and seven members of the public) took part in the interviews, in which four main themes were discussed: a) how participants felt about the genetic engineering of animals as a practice; b) governance of the creation and use of GE animals in research, and whether current guidelines are sufficient; c) the Three Rs (Replacement, Reduction, Refinement) and how they are applied during the creation and use of GE animals in research; and d) whether public opinion should play a greater role in the creation and use of GE animals. Most of the participants felt that the creation and use of GE animals for biomedical research purposes (as opposed to food purposes) is acceptable, provided that tangible human health benefits are gained. However, obstacles to Three Rs implementation were identified, and the participants agreed that more effort should be placed on engaging the public on the use of GE animals in research.

  20. Genetic engineering of cell lines using lentiviral vectors to achieve antibody secretion following encapsulated implantation.

    Science.gov (United States)

    Lathuilière, Aurélien; Bohrmann, Bernd; Kopetzki, Erhard; Schweitzer, Christoph; Jacobsen, Helmut; Moniatte, Marc; Aebischer, Patrick; Schneider, Bernard L

    2014-01-01

    The controlled delivery of antibodies by immunoisolated bioimplants containing genetically engineered cells is an attractive and safe approach for chronic treatments. To reach therapeutic antibody levels there is a need to generate renewable cell lines, which can long-term survive in macroencapsulation devices while maintaining high antibody specific productivity. Here we have developed a dual lentiviral vector strategy for the genetic engineering of cell lines compatible with macroencapsulation, using separate vectors encoding IgG light and heavy chains. We show that IgG expression level can be maximized as a function of vector dose and transgene ratio. This approach allows for the generation of stable populations of IgG-expressing C2C12 mouse myoblasts, and for the subsequent isolation of clones stably secreting high IgG levels. Moreover, we demonstrate that cell transduction using this lentiviral system leads to the production of a functional glycosylated antibody by myogenic cells. Subsequent implantation of antibody-secreting cells in a high-capacity macroencapsulation device enables continuous delivery of recombinant antibodies in the mouse subcutaneous tissue, leading to substantial levels of therapeutic IgG detectable in the plasma.

  1. Genetic Engineering: A Promising Tool to Engender Physiological, Biochemical, and Molecular Stress Resilience in Green Microalgae.

    Science.gov (United States)

    Guihéneuf, Freddy; Khan, Asif; Tran, Lam-Son P

    2016-01-01

    As we march into the 21st century, the prevailing scenario of depleting energy resources, global warming and ever increasing issues of human health and food security will quadruple. In this context, genetic and metabolic engineering of green microalgae complete the quest toward a continuum of environmentally clean fuel and food production. Evolutionarily related, but unlike land plants, microalgae need nominal land or water, and are best described as unicellular autotrophs using light energy to fix atmospheric carbon dioxide (CO2) into algal biomass, mitigating fossil CO2 pollution in the process. Remarkably, a feature innate to most microalgae is synthesis and accumulation of lipids (60-65% of dry weight), carbohydrates and secondary metabolites like pigments and vitamins, especially when grown under abiotic stress conditions. Particularly fruitful, such an application of abiotic stress factors such as nitrogen starvation, salinity, heat shock, etc., can be used in a biorefinery concept for production of multiple valuable products. The focus of this mini-review underlies metabolic reorientation practices and tolerance mechanisms as applied to green microalgae under specific stress stimuli for a sustainable pollution-free future. Moreover, we entail current progress on genetic engineering as a promising tool to grasp adaptive processes for improving strains with potential biotechnological interests.

  2. The morality of socioscientific issues: Construal and resolution of genetic engineering dilemmas

    Science.gov (United States)

    Sadler, Troy D.; Zeidler, Dana L.

    2004-01-01

    The ability to negotiate and resolve socioscientific issues has been posited as integral components of scientific literacy. Although philosophers and science educators have argued that socioscientific issues inherently involve moral and ethical considerations, the ultimate arbiters of morality are individual decision-makers. This study explored the extent to which college students construe genetic engineering issues as moral problems. Twenty college students participated in interviews designed to elicit their ideas, reactions, and feelings regarding a series of gene therapy and cloning scenarios. Qualitative analyses revealed that moral considerations were significant influences on decision-making, indicating a tendency for students to construe genetic engineering issues as moral problems. Students engaged in moral reasoning based on utilitarian analyses of consequences as well as the application of principles. Issue construal was also influenced by affective features such as emotion and intuition. In addition to moral considerations, a series of other factors emerged as important dimensions of socioscientific decision-making. These factors included personal experiences, family biases, background knowledge, and the impact of popular culture. The implications for classroom science instruction and future research are discussed.

  3. Improvements of tolerance to stress conditions by genetic engineering in Saccharomyces cerevisiae during ethanol production.

    Science.gov (United States)

    Doğan, Ayşegül; Demirci, Selami; Aytekin, Ali Özhan; Şahin, Fikrettin

    2014-09-01

    Saccharomyces cerevisiae, industrial yeast isolate, has been of great interest in recent years for fuel ethanol production. The ethanol yield and productivity depend on many inhibitory factors during the fermentation process such as temperature, ethanol, compounds released as the result of pretreatment procedures, and osmotic stress. An ideal strain should be able to grow under different stress conditions occurred at different fermentation steps. Development of tolerant yeast strains can be achieved by reprogramming pathways supporting the ethanol metabolism by regulating the energy balance and detoxicification processes. Complex gene interactions should be solved for an in-depth comprehension of the yeast stress tolerance mechanism. Genetic engineering as a powerful biotechnological tool is required to design new strategies for increasing the ethanol fermentation performance. Upregulation of stress tolerance genes by recombinant DNA technology can be a useful approach to overcome inhibitory situations. This review presents the application of several genetic engineering strategies to increase ethanol yield under different stress conditions including inhibitor tolerance, ethanol tolerance, thermotolerance, and osmotolerance.

  4. Biochemical, genetic, and metabolic engineering strategies to enhance coproduction of 1-propanol and ethanol in engineered Escherichia coli.

    Science.gov (United States)

    Srirangan, Kajan; Liu, Xuejia; Westbrook, Adam; Akawi, Lamees; Pyne, Michael E; Moo-Young, Murray; Chou, C Perry

    2014-11-01

    We recently reported the heterologous production of 1-propanol in Escherichia coli via extended dissimilation of succinate under anaerobic conditions through expression of the endogenous sleeping beauty mutase (Sbm) operon. In the present work, we demonstrate high-level coproduction of 1-propanol and ethanol by developing novel engineered E. coli strains with effective cultivation strategies. Various biochemical, genetic, metabolic, and physiological factors affecting relative levels of acidogenesis and solventogenesis during anaerobic fermentation were investigated. In particular, CPC-PrOH3, a plasmid-free propanogenic E. coli strain derived by activating the Sbm operon on the genome, showed high levels of solventogenesis accounting for up to 85 % of dissimilated carbon. Anaerobic fed-batch cultivation of CPC-PrOH3 with glycerol as the major carbon source produced high titers of nearly 7 g/L 1-propanol and 31 g/L ethanol, implying its potential industrial applicability. The activated Sbm pathway served as an ancillary channel for consuming reducing equivalents upon anaerobic dissimilation of glycerol, resulting in an enhanced glycerol dissimilation and a major metabolic shift from acidogenesis to solventogenesis.

  5. Production of Engineered Fabrics Using Artificial Neural Network-Genetic Algorithm Hybrid Model

    Science.gov (United States)

    Mitra, Ashis; Majumdar, Prabal Kumar; Banerjee, Debamalya

    2015-10-01

    The process of fabric engineering which is generally practised in most of the textile mills is very complicated, repetitive, tedious and time consuming. To eliminate this trial and error approach, a new approach of fabric engineering has been attempted in this work. Data sets of construction parameters [comprising of ends per inch, picks per inch, warp count and weft count] and three fabric properties (namely drape coefficient, air permeability and thermal resistance) of 25 handloom cotton fabrics have been used. The weights and biases of three artificial neural network (ANN) models developed for the prediction of drape coefficient, air permeability and thermal resistance were used to formulate the fitness or objective function and constraints of the optimization problem. The optimization problem was solved using genetic algorithm (GA). In both the fabrics which were attempted for engineering, the target and simulated fabric properties were very close. The GA was able to search the optimum set of fabric construction parameters with reasonably good accuracy except in case of EPI. However, the overall result is encouraging and can be improved further by using larger data sets of handloom fabrics by hybrid ANN-GA model.

  6. Enhancement of lipid production using biochemical, genetic and transcription factor engineering approaches.

    Science.gov (United States)

    Courchesne, Noémie Manuelle Dorval; Parisien, Albert; Wang, Bei; Lan, Christopher Q

    2009-04-20

    This paper compares three possible strategies for enhanced lipid overproduction in microalgae: the biochemical engineering (BE) approaches, the genetic engineering (GE) approaches, and the transcription factor engineering (TFE) approaches. The BE strategy relies on creating a physiological stress such as nutrient-starvation or high salinity to channel metabolic fluxes to lipid accumulation. The GE strategy exploits our understanding to the lipid metabolic pathway, especially the rate-limiting enzymes, to create a channelling of metabolites to lipid biosynthesis by overexpressing one or more key enzymes in recombinant microalgal strains. The TFE strategy is an emerging technology aiming at enhancing the production of a particular metabolite by means of overexpressing TFs regulating the metabolic pathways involved in the accumulation of target metabolites. Currently, BE approaches are the most established in microalgal lipid production. The TFE is a very promising strategy because it may avoid the inhibitive effects of the BE approaches and the limitation of "secondary bottlenecks" as commonly observed in the GE approaches. However, it is still a novel concept to be investigated systematically.

  7. A Pseudomonas putida strain genetically engineered for 1,2,3-trichloropropane bioremediation.

    Science.gov (United States)

    Samin, Ghufrana; Pavlova, Martina; Arif, M Irfan; Postema, Christiaan P; Damborsky, Jiri; Janssen, Dick B

    2014-09-01

    1,2,3-Trichloropropane (TCP) is a toxic compound that is recalcitrant to biodegradation in the environment. Attempts to isolate TCP-degrading organisms using enrichment cultivation have failed. A potential biodegradation pathway starts with hydrolytic dehalogenation to 2,3-dichloro-1-propanol (DCP), followed by oxidative metabolism. To obtain a practically applicable TCP-degrading organism, we introduced an engineered haloalkane dehalogenase with improved TCP degradation activity into the DCP-degrading bacterium Pseudomonas putida MC4. For this purpose, the dehalogenase gene (dhaA31) was cloned behind the constitutive dhlA promoter and was introduced into the genome of strain MC4 using a transposon delivery system. The transposon-located antibiotic resistance marker was subsequently removed using a resolvase step. Growth of the resulting engineered bacterium, P. putida MC4-5222, on TCP was indeed observed, and all organic chlorine was released as chloride. A packed-bed reactor with immobilized cells of strain MC4-5222 degraded >95% of influent TCP (0.33 mM) under continuous-flow conditions, with stoichiometric release of inorganic chloride. The results demonstrate the successful use of a laboratory-evolved dehalogenase and genetic engineering to produce an effective, plasmid-free, and stable whole-cell biocatalyst for the aerobic bioremediation of a recalcitrant chlorinated hydrocarbon. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

  8. Testing the Role of p21-Activated Kinases in Schwannoma Formation Using a Novel Genetically Engineered Murine Model that Closely Phenocopies Human NF2 Disease

    Science.gov (United States)

    2015-06-01

    Kinases in Schwannoma Formation Using a Novel Genetically Engineered Murine Model that Closely Phenocopies Human NF2 Disease The views, opinions and...TITLE AND SUBTITLE Testing the Role of p21-Activated Kinases in Schwannoma Formation Using a Novel Genetically Engineered Murine Model that Closely...1.20 calendar Testing the Role of p21 Activated Kinases in Schwannoma Formation Using a Novel Genetically Engineered Murine Model that Closely

  9. Genetic engineering represents a safe approach for innovations improving nutritional contents of major food crops

    Directory of Open Access Journals (Sweden)

    Werner Arber

    2017-05-01

    Full Text Available About 70 years ago early microbial genetic research revealed that inherited phenotypic traits become determined by DNA filaments composed of 4 different nucleotides that are linearly arranged. In the meantime we know that genes, the determinants of specific life functions, are genomic segments of an average size of about 1000 nucleotides, i.e. a very small part of a genome. Fundamental insights into the structures and functions of selected genes can be reached by sorting out the relevant short DNA segment, splicing this fragment into a natural gene vector such as a viral genome or a fertility plasmid. This allows the researchers to transfer the genetic hybrid into an appropriate host cell in order to produce many copies that can then serve for functional and structural analysis. This research approach became efficient in the 1970s. On the request of involved researchers, safety guidelines became proposed 1975 at the Asilomar Conference on Recombinant DNA (Berg, Baltimore, Brenner, Roblin, & Singer, 1975, then generally introduced and still largely followed nowadays. Carefully carried out genetic engineering by horizontally transferring a selected and functionally well known DNA segment into the genome of another organism has in many published biosafety investigations never shown any unexpected harmful effect. We will present below selected examples of research contributions enabling innovations for the benefit of human life conditions.

  10. Space mutagenesis of genetically engineered bacteria expressing recombinant human interferon α1b and screening of higher yielding strains.

    Science.gov (United States)

    Wang, Junfeng; Liu, Changting; Liu, Jinyi; Fang, Xiangqun; Xu, Chen; Guo, Yinghua; Chang, De; Su, Longxiang

    2014-03-01

    The aim of this study was to investigate the space mutagenesis of genetically engineered bacteria expressing recombinant human interferon α1b. The genetically engineered bacteria expressing the recombinant interferon α1b were sent into outer space on the Chinese Shenzhou VIII spacecraft. After the 17 day space flight, mutant strains that highly expressed the target gene were identified. After a series of screening of spaceflight-treated bacteria and the quantitative comparison of the mutant strains and original strain, we found five strains that showed a significantly higher production of target proteins, compared with the original strain. Our results support the notion that the outer space environment has unique effects on the mutation breeding of microorganisms, including genetically engineered strains. Mutant strains that highly express the target protein could be obtained through spaceflight-induced mutagenesis.

  11. Biochemical and Genetic Engineering of Diatoms for Polyunsaturated Fatty Acid Biosynthesis

    Directory of Open Access Journals (Sweden)

    Hong-Ye Li

    2014-01-01

    Full Text Available The role of diatoms as a source of bioactive compounds has been recently explored. Diatom cells store a high amount of fatty acids, especially certain polyunsaturated fatty acids (PUFAs. However, many aspects of diatom metabolism and the production of PUFAs remain unclear. This review describes a number of technical strategies, such as modulation of environmental factors (temperature, light, chemical composition of culture medium and culture methods, to influence the content of PUFAs in diatoms. Genetic engineering, a newly emerging field, also plays an important role in controlling the synthesis of fatty acids in marine microalgae. Several key points in the biosynthetic pathway of PUFAs in diatoms as well as recent progresses are also a critical part and are summarized here.

  12. Heritable multiplex genetic engineering in rats using CRISPR/Cas9.

    Directory of Open Access Journals (Sweden)

    Yuanwu Ma

    Full Text Available The CRISPR/Cas9 system has been proven to be an efficient gene-editing tool for genome modification of cells and organisms. Multiplex genetic engineering in rat holds a bright future for the study of complex disease. Here, we show that this system enables the simultaneous disruption of four genes (ApoE, B2m, Prf1, and Prkdc in rats in one-step, by co-injection of Cas9 mRNA and sgRNAs into fertilized eggs. We further observed the gene modifications are germline transmittable, and confirmed the off-target mutagenesis and mosaicism are rarely detected by comprehensive analysis. Thus, the CRISPR/Cas9 system makes it possible to efficiently and reliably generate gene knock-out rats.

  13. Can we build it better? Using BAC genetics to engineer more effective cytomegalovirus vaccines.

    Science.gov (United States)

    Schleiss, Mark R

    2010-12-01

    The magnitude and durability of immunity to human cytomegalovirus (HCMV) following natural infection is compromised by the presence of immune modulation genes that appear to promote evasion of host clearance mechanisms. Since immunity to HCMV offers limited protection, rational design of effective vaccines has been challenging. In this issue of the JCI, Slavuljica and colleagues employ techniques to genetically modify the highly related mouse CMV (MCMV), in the process generating a virus that was rapidly cleared by NK cells. The virus functioned as a safe and highly effective vaccine. Demonstration of the ability to engineer a safe and highly effective live virus vaccine in a relevant rodent model of CMV infection may open the door to clinical trials of safer and more immunogenic HCMV vaccines.

  14. Overview of Genetically Engineered Mouse Models of Breast Cancer Used in Translational Biology and Drug Development.

    Science.gov (United States)

    Greenow, Kirsty R; Smalley, Matthew J

    2015-01-01

    Breast cancer is a heterogeneous condition with no single standard of treatment and no definitive method for determining whether a tumor will respond to therapy. The development of murine models that faithfully mimic specific human breast cancer subtypes is critical for the development of patient-specific treatments. While the artificial nature of traditional in vivo xenograft models used to characterize novel anticancer treatments has limited clinical predictive value, the development of genetically engineered mouse models (GEMMs) makes it possible to study the therapeutic responses in an intact microenvironment. GEMMs have proven to be an experimentally tractable platform for evaluating the efficacy of novel therapeutic combinations and for defining the mechanisms of acquired resistance. Described in this overview are several of the more popular breast cancer GEMMs, including details on their value in elucidating the molecular mechanisms of this disorder.

  15. Biochemical and genetic engineering strategies to enhance hydrogen production in photosynthetic algae and cyanobacteria.

    Science.gov (United States)

    Srirangan, Kajan; Pyne, Michael E; Perry Chou, C

    2011-09-01

    As an energy carrier, hydrogen gas is a promising substitute to carbonaceous fuels owing to its superb conversion efficiency, non-polluting nature, and high energy content. At present, hydrogen is predominately synthesized via chemical reformation of fossil fuels. While various biological methods have been extensively explored, none of them is justified as economically feasible. A sustainable platform for biological production of hydrogen will certainly impact the biofuel market. Among a selection of biological systems, algae and cyanobacteria have garnered major interests as potential cell factories for hydrogen production. In conjunction with photosynthesis, these organisms utilize inexpensive inorganic substrates and solar energy for simultaneous biosynthesis and hydrogen evolution. However, the hydrogen yield associated with these organisms remains far too low to compete with the existing chemical systems. This article reviews recent advances of biochemical, bioprocess, and genetic engineering strategies in circumventing technological limitations to hopefully improve the applicative potential of these photosynthetic hydrogen production systems. Copyright © 2011 Elsevier Ltd. All rights reserved.

  16. Genetically engineered mouse models to evaluate the role of Wnt secretion in bone development and homeostasis.

    Science.gov (United States)

    Williams, Bart O

    2016-03-01

    Alterations in components of the Wnt signaling pathway are associated with altered bone development and homeostasis in several human diseases. We created genetically engineered mouse models (GEMMs) that mimic the cellular defect associated with the Porcupine mutations in patients with Goltz Syndrome/Focal Dermal Hypoplasia. These GEMMs were established by utilizing mice containing a conditionally inactivatable allele of Wntless/GPR177 (a gene encoding a protein required for the transport of Porcupine-modified ligand to the plasma membrane for secretion). We crossed this strain to another which drives cre-mediated gene deletion in mature osteoblasts (Osteocalcin-cre) resulted in mice lacking the ability to secrete Wnt ligands in this cell type. These mice displayed severely reduced bone mass and provide a model to understand the effects of disrupting the ability to secrete Wnt ligands on the skeletal system.

  17. Analysis of mouse model pathology: a primer for studying the anatomic pathology of genetically engineered mice.

    Science.gov (United States)

    Cardiff, Robert D; Miller, Claramae H; Munn, Robert J

    2014-06-02

    This primer of pathology is intended to introduce investigators to the structure (morphology) of cancer with an emphasis on genetically engineered mouse (GEM) models (GEMMs). We emphasize the necessity of using the entire biological context for the interpretation of anatomic pathology. Because the primary investigator is responsible for almost all of the information and procedures leading up to microscopic examination, they should also be responsible for documentation of experiments so that the microscopic interpretation can be rendered in context of the biology. The steps involved in this process are outlined, discussed, and illustrated. Because GEMMs are unique experimental subjects, some of the more common pitfalls are discussed. Many of these errors can be avoided with attention to detail and continuous quality assurance.

  18. Biochemical and genetic engineering of diatoms for polyunsaturated fatty acid biosynthesis.

    Science.gov (United States)

    Li, Hong-Ye; Lu, Yang; Zheng, Jian-Wei; Yang, Wei-Dong; Liu, Jie-Sheng

    2014-01-07

    The role of diatoms as a source of bioactive compounds has been recently explored. Diatom cells store a high amount of fatty acids, especially certain polyunsaturated fatty acids (PUFAs). However, many aspects of diatom metabolism and the production of PUFAs remain unclear. This review describes a number of technical strategies, such as modulation of environmental factors (temperature, light, chemical composition of culture medium) and culture methods, to influence the content of PUFAs in diatoms. Genetic engineering, a newly emerging field, also plays an important role in controlling the synthesis of fatty acids in marine microalgae. Several key points in the biosynthetic pathway of PUFAs in diatoms as well as recent progresses are also a critical part and are summarized here.

  19. Biosynthesis of 2-phenylethanol from glucose with genetically engineered Kluyveromyces marxianus.

    Science.gov (United States)

    Kim, Tae-Yeon; Lee, Sang-Woo; Oh, Min-Kyu

    2014-01-01

    2-Phenylethanol (2-PE) is an aromatic alcohol with a rose scent, which is used in the cosmetics, fragrance and food industries. 2-PE is produced in a few yeast strains by Ehrlich pathway. In this study, Kluyveromyces marxianus was genetically engineered for overproduction of 2-PE from glucose. About 1.0g/L of 2-PE was produced by overexpressing phenylpyruvate decarboxylase (ARO10) and alcohol dehydrogenase (ADH2) genes of Saccharomyces cerevisiae. A similar level of 2-PE was also produced from evolved K. marxianus, which was resistant to the phenylalanine analog, p-fluorophenylalanine. aroG(fbr) from Klebsiella pneumoniae encoding a feedback resistant mutant of 3-deoxy-D-arabino-heptulosonate-7-phosphate (DHAP) synthase was overexpressed in the evolved K. marxianus. Finally, 1.3g/L of 2-PE was produced from 20g/L glucose without addition of phenylalanine in the medium.

  20. The establishment of genetically engineered canola populations in the U.S.

    Directory of Open Access Journals (Sweden)

    Meredith G Schafer

    Full Text Available Concerns regarding the commercial release of genetically engineered (GE crops include naturalization, introgression to sexually compatible relatives and the transfer of beneficial traits to native and weedy species through hybridization. To date there have been few documented reports of escape leading some researchers to question the environmental risks of biotech products. In this study we conducted a systematic roadside survey of canola (Brassica napus populations growing outside of cultivation in North Dakota, USA, the dominant canola growing region in the U.S. We document the presence of two escaped, transgenic genotypes, as well as non-GE canola, and provide evidence of novel combinations of transgenic forms in the wild. Our results demonstrate that feral populations are large and widespread. Moreover, flowering times of escaped populations, as well as the fertile condition of the majority of collections suggest that these populations are established and persistent outside of cultivation.

  1. Genetic engineering of modular PKSs: from combinatorial biosynthesis to synthetic biology.

    Science.gov (United States)

    Weissman, Kira J

    2016-02-01

    Multienzyme polyketide synthases (PKSs) are molecular-scale assembly lines which construct complex natural products in bacteria. The underlying modular architecture of these gigantic catalysts inspired, from the moment of their discovery, attempts to modify them by genetic engineering to produce analogues of predictable structure. These efforts have resulted in hundreds of metabolites new to nature, as detailed in this review. However, in the face of many failures, the heady days of imagining the possibilities for a truly 'combinatorial biosynthesis' of polyketides have faded. It is now more appropriate to talk about 'PKS synthetic biology' with its more modest goals of delivering specific derivatives of known structure in combination with and as a complement to synthetic chemistry approaches. The reasons for these failures will be discussed in terms of our growing understanding of the three-dimensional architectures and mechanisms of these systems. Finally, some thoughts on the future of the field will be presented.

  2. Genetic Engineering of Cyanobacteria to Enhance Biohydrogen Production from Sunlight and Water

    Energy Technology Data Exchange (ETDEWEB)

    Masukawa, Hajime (Research Inst. for Photobiological Hydrogen Production, Kanagawa Univ., Hiratsuka, Kanagawa (Japan); PRESTO, Japan Science and Technology Agency, Kawaguchi, Saitama (Japan)), E-mail: wtk-0488gg@kanagawa-u.ac.jp; Kitashima, Masaharu (Research Inst. for Integrated Science, Kanagawa Univ., Hiratsuka, Kanagawa (Japan)); Inoue, Kazuhito (Dept. of Biological Sciences, Kanagawa Univ., Hiratsuka, Kanagawa (Japan)); Sakurai, Hidehiro (Research Inst. for Photobiological Hydrogen Production, Kanagawa Univ., Hiratsuka, Kanagawa (Japan)); Hausinger, Robert P. (Dept. of Microbiology and Molecular Genetics, 2215 Biomedical Physical Sciences, Michigan State Univ., East Lansing (United States))

    2012-03-15

    To mitigate global warming caused by burning fossil fuels, a renewable energy source available in large quantity is urgently required. We are propoi large-scale photobiological H{sub 2} production by mariculture-raised cyanobacteria where the microbes capture part of the huge amount of solar energy received on earth's surface and use water as the source of electrons to reduce protons. The H{sub 2} production system is based on photosynthetic and nitrogenase activities of cyanobacteria, using uptake hydrogenase mutants that can accumulate H{sub 2} for extended periods even in the presence of evolved O{sub 2}. This review summarizes our efforts to improve the rate of photobiological H{sub 2} production through genetic engineering. The challenges yet to be overcome to further increase the conversion efficiency of solar energy to H{sub 2} also are discussed

  3. Over production of lignocellulosic enzymes of Coriolus versicolor by genetic engineering methodology. Final report

    Energy Technology Data Exchange (ETDEWEB)

    Williams, A.L.

    1998-07-01

    The project seeks to understand the biological and chemical processes involved in the secretion of the enzyme polyphenol oxidase (PPO) by the hyphae, the basic unit of the filamentous fungus Coriolus versicolor. These studies are made to determine rational strategies for enhanced secretion of PPO, both with the use of recombinant DNA techniques and without. This effort focuses on recombinant DNA techniques to enhance enzyme production. The major thrust of this project was two-fold: to mass produce C. versicolor tyrosinase (polyphenol oxidase) by genetic engineering as well as cultural manipulations; and to utilize PPO as a biocatalyst in the processing of lignocellulose as a renewable energy resource. In this study, the assessment of genomic and cDNA recombinant clones with regards to the overproduction of PPO continued. Further, immunocytochemical techniques were employed to assess the mechanism(s) involved in the secretion of PPO by the hyphae. Also, factors influencing PPO secretion were examined.

  4. Lentivectors encoding immunosuppressive proteins genetically engineer pancreatic beta-cells to correct diabetes in allogeneic mice.

    Science.gov (United States)

    Kojaoghlanian, T; Joseph, A; Follenzi, A; Zheng, J H; Leiser, M; Fleischer, N; Horwitz, M S; DiLorenzo, T P; Goldstein, H

    2009-03-01

    The effectiveness of genetic engineering with lentivectors to protect transplanted cells from allogeneic rejection was examined using, as a model, type 1 diabetes treatment with beta-cell transplantation, whose widespread use has been limited by the requirement for sustained immunosuppressive treatment to prevent graft rejection. We examined whether lentivectors expressing select immunosuppressive proteins encoded by the adenoviral genome early region 3 (AdE3) would protect transplanted beta-cells from an alloimmune attack. The insulin-producing beta-cell line beta TC-tet (C3HeB/FeJ-derived) was transduced with lentiviruses encoding the AdE3 proteins gp19K and RID alpha/beta. The efficiency of lentiviral transduction of beta TC-tet cells exceeded 85%. Lentivector expression of gp19K decreased surface class I major histocompatibility complex expression by over 90%, whereas RID alpha/beta expression inhibited cytokine-induced Fas upregulation by over 75%. beta TC-tet cells transduced with gp19K and RID alpha/beta lentivectors, but not with a control lentivector, provided prolonged correction of hyperglycemia after transplantation into diabetic BALB/c severe combined immunodeficient mice reconstituted with allogeneic immune effector cells or into diabetic allogeneic BALB/c mice. Thus, genetic engineering of beta-cells using gp19K- and RID alpha/beta-expressing lentiviral vectors may provide an alternative that has the potential to eliminate or reduce treatment with the potent immunosuppressive agents necessary at present for prolonged engraftment with transplanted islets.

  5. Development of New Modular Genetic Tools for Engineering the Halophilic Archaeon Halobacterium salinarum.

    Directory of Open Access Journals (Sweden)

    Rafael Silva-Rocha

    Full Text Available Our ability to genetically manipulate living organisms is usually constrained by the efficiency of the genetic tools available for the system of interest. In this report, we present the design, construction and characterization of a set of four new modular vectors, the pHsal series, for engineering Halobacterium salinarum, a model halophilic archaeon widely used in systems biology studies. The pHsal shuttle vectors are organized in four modules: (i the E. coli's specific part, containing a ColE1 origin of replication and an ampicillin resistance marker, (ii the resistance marker and (iii the replication origin, which are specific to H. salinarum and (iv the cargo, which will carry a sequence of interest cloned in a multiple cloning site, flanked by universal M13 primers. Each module was constructed using only minimal functional elements that were sequence edited to eliminate redundant restriction sites useful for cloning. This optimization process allowed the construction of vectors with reduced sizes compared to currently available platforms and expanded multiple cloning sites. Additionally, the strong constitutive promoter of the fer2 gene was sequence optimized and incorporated into the platform to allow high-level expression of heterologous genes in H. salinarum. The system also includes a new minimal suicide vector for the generation of knockouts and/or the incorporation of chromosomal tags, as well as a vector for promoter probing using a GFP gene as reporter. This new set of optimized vectors should strongly facilitate the engineering of H. salinarum and similar strategies could be implemented for other archaea.

  6. Examining strategies to facilitate vitamin B1 biofortification of plants by genetic engineering

    Directory of Open Access Journals (Sweden)

    Lucille ePourcel

    2013-05-01

    Full Text Available Thiamin (vitamin B1 is made by plants and microorganisms but is an essential micronutrient in the human diet. All organisms require it as a cofactor in its form as thiamin pyrophosphate (TPP for the activity of key enzymes of central metabolism. In humans, deficiency is widespread particularly in populations where polished rice is a major component of the diet. Considerable progress has been made on the elucidation of the biosynthesis pathway within the last few years enabling concrete strategies for biofortification purposes to be devised, with a particular focus here on genetic engineering. Furthermore, the vitamin has been shown to play a role in both abiotic and biotic stress responses. The precursors for de novo biosynthesis of thiamin differ between microorganisms and plants. Bacteria use intermediates derived from purine and isoprenoid biosynthesis, whereas the pathway in yeast involves the use of compounds from the vitamin B3 and B6 groups. Plants on the other hand use a combination of the bacterial and yeast pathways and there is subcellular partitioning of the biosynthesis steps. Specifically, thiamin biosynthesis occurs in the chloroplast of plants through the separate formation of the pyrimidine and thiazole moieties, which are then coupled to form thiamin monophosphate (TMP. Phosphorylation of thiamin to form TPP occurs in the cytosol. Therefore, thiamin (or TMP must be exported from the chloroplast to the cytosol for the latter step to be executed. The regulation of biosynthesis is mediated through riboswitches, where binding of the product TPP to the pre-mRNA of a biosynthetic gene modulates expression. Here we examine and hypothesize on genetic engineering approaches attempting to increase the thiamin content employing knowledge gained with the model plant Arabidopsis thaliana. We will discuss the regulatory steps that need to be taken into consideration and can be used a prerequisite for devising such strategies in crop plants.

  7. Release of tissue inhibitor of metalloproteinase-2 from alginate microcapsule encapsulating genetically engineered cells

    Directory of Open Access Journals (Sweden)

    Kim YS

    2013-11-01

    Full Text Available Yeon Seong Kim,1,* Young-Il Jeong,2,* Shu-Guang Jin,2 Jian Pei,2 Min Wen,2 In-Young Kim,1 Kyung-Sub Moon,1 Tae-Young Jung,1 Hyang-Hwa Ryu2, Shin Jung1–3 1Department of Neurosurgery, 2Brain Tumor Research Laboratory, 3Chonnam National University Research Institute of Medical Sciences, Chonnam National University Hwasun Hospital and Medical School, Jeollanam-do, Korea *These authors contributed equally to this work Background: In this study, 293T cells were genetically engineered to secrete tissue inhibitor of metalloproteinase-2 (TIMP2 and encapsulated into alginate microcapsules to continuously release TIMP2 protein. Methods: The anti-invasive potential of the microcapsules was studied in vitro using brain tumor cells. The TIMP2 gene was transfected to 293T cells, and genetically engineered 293TIMP2 cells were encapsulated into alginate microcapsules. Release of TIMP2 protein was detected with Western blot analysis and the anti-invasive potential against U87MG cells was tested using gelatin zymography and a Matrigel assay. Results: Cell viability within the alginate microcapsules was maintained at a cell density of 5 × 106. Because polycationic polymers are helpful for maintaining the mechanical strength of microcapsules with good cell viability, the alginate microcapsules were reinforced with chitosan (0.1% w/v. Expression of TIMP2 protein in cell lysates and secretion of TIMP2 into the conditioned medium was confirmed by Western blot analysis. Alginate microcapsules encapsulating 293TIMP2 cells released TIMP2 protein into the medium efficiently, where the TIMP2 protein participated in degradation of the matrix metalloproteinase-2 enzyme and inhibited invasion of U87MG cells. Conclusion: Alginate microcapsules encapsulating 293TIMP2 cells are promising candidates for anti-invasive treatment of glioma. Keywords: 293T cells, tissue inhibitor of metalloproteinase-2, alginate microcapsule, therapeutic protein

  8. Genetic engineering of novel flower colour by suppression of anthocyanin modification genes in gentian.

    Science.gov (United States)

    Nakatsuka, Takashi; Mishiba, Kei-ichiro; Kubota, Akiko; Abe, Yoshiko; Yamamura, Saburo; Nakamura, Noriko; Tanaka, Yoshikazu; Nishihara, Masahiro

    2010-02-15

    Ornamental gentian plants have vivid-blue flowers. The main factor contributing to the flower colour is the accumulation of a polyacylated delphinidin 'gentiodelphin' in their petals. Although in vitro studies proposed that acylation plays an important role in the stability and development of gentian blue colour, the in vivo stability of the polyacylated anthocyanin was not clearly demonstrated. Thus, to reveal the importance of anthocyanin modification, especially acylation, and to engineer new colours of gentian flowers, we used chimeric RNAi technology to produce transgenic gentian plants with downregulated anthocyanin 5,3'-aromatic acyltransferase (5/3'AT) and flavonoid 3',5'-hydroxylase (F3'5'H) activities, which are both essential enzymes for gentiodelphin biosynthesis. Two lines of flower colour-modified plants were obtained from fifteen transgenic gentian plants. Clone no. 1 exhibited a lilac flower colour and clone no. 15 exhibited pale-blue flowers. RNA gel blot analysis confirmed that both transgenic lines had markedly suppressed 5/3'AT transcripts, whereas clone no. 15 had fewer F3'5'H transcripts than clone no. 1 and untransformed control plants. HPLC analysis of anthocyanin compositions showed that downregulation of the 5/3'AT gene led to increased accumulation of non-acylated anthocyanins, as expected. These results demonstrated that genetic engineering to reduce the accumulation of polyacylated anthocyanins could cause modulations of flower colour.

  9. A prototype stable RNA identification cassette for monitoring plasmids of genetically engineered microorganisms

    Science.gov (United States)

    Hedenstierna, K. O.; Lee, Y. H.; Yang, Y.; Fox, G. E.

    1993-01-01

    A prototype stable RNA identification cassette for monitoring genetically engineered plasmids carried by strains of Escherichia coli has been developed. The cassette consists of a Vibrio proteolyticus 5S ribosomal RNA (rRNA) gene surrounded by promoters and terminators from the rrnB operon of Escherischia coli. The identifier RNA is expressed and successfully processed so that approximately 30% of the 5S rRNA isolated from either whole cells or 70S ribosomes is of the V. proteolyticus type. Cells carrying the identifier are readily detectable by hybridization. Accurate measurements show that the identification cassette has little effect on fitness compared to a strain containing an analogous plasmid carrying wild type E. coli 5S rRNA, and the V. proteolyticus 5S rRNA gene is not inactivated after prolonged growth. These results demonstrate the feasibility of developing small standardized identification cassettes that can utilize already existing highly sensitive rRNA detection methods. Cassettes of this type could in principle be incorporated into either the engineered regions of recombinant plasmids or their hosts.

  10. Genetic engineering and metabolite profiling for overproduction of polyhydroxybutyrate in cyanobacteria.

    Science.gov (United States)

    Hondo, Sayaka; Takahashi, Masatoshi; Osanai, Takashi; Matsuda, Mami; Hasunuma, Tomohisa; Tazuke, Akio; Nakahira, Yoichi; Chohnan, Shigeru; Hasegawa, Morifumi; Asayama, Munehiko

    2015-11-01

    Genetic engineering and metabolite profiling for the overproduction of polyhydroxybutyrate (PHB), which is a carbon material in biodegradable plastics, were examined in the unicellular cyanobacterium Synechocystis sp. PCC 6803. Transconjugants harboring cyanobacterial expression vectors that carried the pha genes for PHB biosynthesis were constructed. The overproduction of PHB by the engineering cells was confirmed through microscopic observations using Nile red, transmission electron microscopy (TEM), or nuclear magnetic resonance (NMR). We successfully recovered PHB from transconjugants prepared from nitrogen-depleted medium without sugar supplementation in which PHB reached approximately 7% (w/w) of the dry cell weight, showing a value of 12-fold higher productivity in the transconjugant than that in the control strain. We also measured the intracellular levels of acetyl-CoA, acetoacetyl-CoA, and 3-hydroxybutyryl-CoA (3HB-CoA), which are intermediate products for PHB. The results obtained indicated that these products were absent or at markedly low levels when cells were subjected to the steady-state growth phase of cultivation under nitrogen depletion for the overproduction of bioplastics. Based on these results, efficient factors were discussed for the overproduction of PHB in recombinant cyanobacteria.

  11. Genetically engineered virus-resistant plants in developing countries: current status and future prospects.

    Science.gov (United States)

    Reddy, D V R; Sudarshana, M R; Fuchs, M; Rao, N C; Thottappilly, G

    2009-01-01

    Plant viruses cause severe crop losses worldwide. Conventional control strategies, such as cultural methods and biocide applications against arthropod, nematode, and plasmodiophorid vectors, have limited success at mitigating the impact of plant viruses. Planting resistant cultivars is the most effective and economical way to control plant virus diseases. Natural sources of resistance have been exploited extensively to develop virus-resistant plants by conventional breeding. Non-conventional methods have also been used successfully to confer virus resistance by transferring primarily virus-derived genes, including viral coat protein, replicase, movement protein, defective interfering RNA, non-coding RNA sequences, and protease, into susceptible plants. Non-viral genes (R genes, microRNAs, ribosome-inactivating proteins, protease inhibitors, dsRNAse, RNA modifying enzymes, and scFvs) have also been used successfully to engineer resistance to viruses in plants. Very few genetically engineered (GE) virus resistant (VR) crops have been released for cultivation and none is available yet in developing countries. However, a number of economically important GEVR crops, transformed with viral genes are of great interest in developing countries. The major issues confronting the production and deregulation of GEVR crops in developing countries are primarily socio-economic and related to intellectual property rights, biosafety regulatory frameworks, expenditure to generate GE crops and opposition by non-governmental activists. Suggestions for satisfactory resolution of these factors, presumably leading to field tests and deregulation of GEVR crops in developing countries, are given.

  12. A genetic replacement system for selection-based engineering of essential proteins

    Directory of Open Access Journals (Sweden)

    Billerbeck Sonja

    2012-08-01

    Full Text Available Abstract Background Essential genes represent the core of biological functions required for viability. Molecular understanding of essentiality as well as design of synthetic cellular systems includes the engineering of essential proteins. An impediment to this effort is the lack of growth-based selection systems suitable for directed evolution approaches. Results We established a simple strategy for genetic replacement of an essential gene by a (library of variant(s during a transformation. The system was validated using three different essential genes and plasmid combinations and it reproducibly shows transformation efficiencies on the order of 107 transformants per microgram of DNA without any identifiable false positives. This allowed for reliable recovery of functional variants out of at least a 105-fold excess of non-functional variants. This outperformed selection in conventional bleach-out strains by at least two orders of magnitude, where recombination between functional and non-functional variants interfered with reliable recovery even in recA negative strains. Conclusions We propose that this selection system is extremely suitable for evaluating large libraries of engineered essential proteins resulting in the reliable isolation of functional variants in a clean strain background which can readily be used for in vivo applications as well as expression and purification for use in in vitro studies.

  13. Site-specific genetic engineering of the Anopheles gambiae Y chromosome.

    Science.gov (United States)

    Bernardini, Federica; Galizi, Roberto; Menichelli, Miriam; Papathanos, Philippos-Aris; Dritsou, Vicky; Marois, Eric; Crisanti, Andrea; Windbichler, Nikolai

    2014-05-27

    Despite its function in sex determination and its role in driving genome evolution, the Y chromosome remains poorly understood in most species. Y chromosomes are gene-poor, repeat-rich and largely heterochromatic and therefore represent a difficult target for genetic engineering. The Y chromosome of the human malaria vector Anopheles gambiae appears to be involved in sex determination although very little is known about both its structure and function. Here, we characterize a transgenic strain of this mosquito species, obtained by transposon-mediated integration of a transgene construct onto the Y chromosome. Using meganuclease-induced homologous repair we introduce a site-specific recombination signal onto the Y chromosome and show that the resulting docking line can be used for secondary integration. To demonstrate its utility, we study the activity of a germ-line-specific promoter when located on the Y chromosome. We also show that Y-linked fluorescent transgenes allow automated sex separation of this important vector species, providing the means to generate large single-sex populations. Our findings will aid studies of sex chromosome function and enable the development of male-exclusive genetic traits for vector control.

  14. On recent advances in human engineering Provocative trends in embryology, genetics, and regenerative medicine.

    Science.gov (United States)

    Anton, Roman

    2016-01-01

    Advances in embryology, genetics, and regenerative medicine regularly attract attention from scientists, scholars, journalists, and policymakers, yet implications of these advances may be broader than commonly supposed. Laboratories culturing human embryos, editing human genes, and creating human-animal chimeras have been working along lines that are now becoming intertwined. Embryogenic methods are weaving traditional in vivo and in vitro distinctions into a new "in vivitro" (in life in glass) fabric. These and other methods known to be in use or thought to be in development promise soon to bring society to startling choices and discomfiting predicaments, all in a global effort to supply reliably rejuvenating stem cells, to grow immunologically non-provocative replacement organs, and to prevent, treat, cure, or even someday eradicate diseases having genetic or epigenetic mechanisms. With humanity's human-engineering era now begun, procedural prohibitions, funding restrictions, institutional controls, and transparency rules are proving ineffective, and business incentives are migrating into the most basic life-sciences inquiries, wherein lie huge biomedical potentials and bioethical risks. Rights, health, and heritage are coming into play with bioethical presumptions and formal protections urgently needing reassessment.

  15. Rifampicin-resistance, rpoB polymorphism and RNA polymerase genetic engineering.

    Science.gov (United States)

    Alifano, Pietro; Palumbo, Carla; Pasanisi, Daniela; Talà, Adelfia

    2015-05-20

    Following its introduction in 1967, rifampicin has become a mainstay of therapy in the treatment of tuberculosis, leprosy and many other widespread diseases. Its potent antibacterial activity is due to specific inhibition of bacterial RNA polymerase. However, resistance to rifampicin was reported shortly after its introduction in the medical practice. Studies in the model organism Escherichia coli helped to define the molecular mechanism of rifampicin-resistance demonstrating that resistance is mostly due to chromosomal mutations in rpoB gene encoding the RNA polymerase β chain. These studies also revealed the amazing potential of the molecular genetics to elucidate the structure-function relationships in bacterial RNA polymerase. The scope of this paper is to illustrate how rifampicin-resistance has been recently exploited to better understand the regulatory mechanisms that control bacterial cell physiology and virulence, and how this information has been used to maneuver, on a global scale, gene expression in bacteria of industrial interest. In particular, we reviewed recent literature regarding: (i) the effects of rpoB mutations conferring rifampicin-resistance on transcription dynamics, bacterial fitness, physiology, metabolism and virulence; (ii) the occurrence in nature of "mutant-type" or duplicated rifampicin-resistant RNA polymerases; and (iii) the RNA polymerase genetic engineering method for strain improvement and drug discovery.

  16. Frontiers of torenia research: innovative ornamental traits and study of ecological interaction networks through genetic engineering.

    Science.gov (United States)

    Nishihara, Masahiro; Shimoda, Takeshi; Nakatsuka, Takashi; Arimura, Gen-Ichiro

    2013-06-26

    Advances in research in the past few years on the ornamental plant torenia (Torenia spps.) have made it notable as a model plant on the frontier of genetic engineering aimed at studying ornamental characteristics and pest control in horticultural ecosystems. The remarkable advantage of torenia over other ornamental plant species is the availability of an easy and high-efficiency transformation system for it. Unfortunately, most of the current torenia research is still not very widespread, because this species has not become prominent as an alternative to other successful model plants such as Arabidopsis, snapdragon and petunia. However, nowadays, a more global view using not only a few selected models but also several additional species are required for creating innovative ornamental traits and studying horticultural ecosystems. We therefore introduce and discuss recent research on torenia, the family Scrophulariaceae, for secondary metabolite bioengineering, in which global insights into horticulture, agriculture and ecology have been advanced. Floral traits, in torenia particularly floral color, have been extensively studied by manipulating the flavonoid biosynthetic pathways in flower organs. Plant aroma, including volatile terpenoids, has also been genetically modulated in order to understand the complicated nature of multi-trophic interactions that affect the behavior of predators and pollinators in the ecosystem. Torenia would accordingly be of great use for investigating both the variation in ornamental plants and the infochemical-mediated interactions with arthropods.

  17. Pathway engineering for phenolic acid accumulations in Salvia miltiorrhiza by combinational genetic manipulation.

    Science.gov (United States)

    Zhang, Yuan; Yan, Ya-Ping; Wu, Yu-Cui; Hua, Wen-Ping; Chen, Chen; Ge, Qian; Wang, Zhe-Zhi

    2014-01-01

    To produce beneficial phenolic acids for medical and commercial purposes, researchers are interested in improving the normally low levels of salvianolic acid B (Sal B) produced by Salvia miltiorrhiza. Here, we present a strategy of combinational genetic manipulation to enrich the precursors available for Sal B biosynthesis. This approach, involving the lignin pathway, requires simultaneous, ectopic expression of an Arabidopsis Production of Anthocyanin Pigment 1 transcription factor (AtPAP1) plus co-suppression of two endogenous, key enzyme genes: cinnamoyl-CoA reductase (SmCCR) and caffeic acid O-methyltransferase (SmCOMT). Compared with the untransformed control, we achieved a greater accumulation of Sal B (up to 3-fold higher) along with a reduced lignin concentration. This high-Sal B phenotype was stable in roots during vegetative growth and was closely correlated with increased antioxidant capacity for the corresponding plant extracts. Although no outward change in phenotype was apparent, we characterized the molecular phenotype through integrated analysis of transcriptome and metabolome profiling. Our results demonstrated the far-reaching consequences of phenolic pathway perturbations on carbohydrate metabolism, respiration, photo-respiration, and stress responses. This report is the first to describe the production of valuable end products through combinational genetic manipulation in S. miltiorrhiza plants. Our strategy will be effective in efforts to metabolically engineer multi-branch pathway(s), such as the phenylpropanoid pathway, in economically significant medicinal plants.

  18. Synthesis of magnetite nanoparticles for bio- and nanotechnology: genetic engineering and biomimetics of bacterial magnetosomes.

    Science.gov (United States)

    Lang, Claus; Schüler, Dirk; Faivre, Damien

    2007-02-12

    Magnetotactic bacteria (MTB) have the ability to navigate along the Earth's magnetic field. This so-called magnetotaxis is a result of the presence of magnetosomes, organelles which comprise nanometer-sized intracellular crystals of magnetite (Fe(3)O(4)) enveloped by a membrane. Because of their unique characteristics, magnetosomes have a high potential for nano- and biotechnological applications, which require a specifically designed particle surface. The functionalization of magnetosomes is possible either by chemical modification of purified particles or by genetic engineering of magnetosome membrane proteins. The second approach is potentially superior to chemical approaches as a large variety of biological functions such as protein tags, fluorophores, and enzymes may be directly incorporated in a site-specific manner during magnetosome biomineralization. An alternative to the bacterial production of magnetosomes are biomimetic approaches, which aim to mimic the bacterial biomineralization pathway in vitro. In MTB a number of magnetosome proteins with putative functions in the biomineralization of the nanoparticles have been identified by genetic and biochemical approaches. The initial results obtained by several groups indicate that some of these proteins have an impact on nanomagnetite properties in vitro. In this article the key features of magnetosomes are discussed, an overview of their potential applications are given, and different strategies are proposed for the functionalization of magnetosome particles and for the biomimetism of their biomineralization pathway.

  19. Genetically Engineered Corn Rootworm Resistance: Potential for Reduction of Human Health Effects From Pesticides

    Institute of Scientific and Technical Information of China (English)

    2003-01-01

    Objective and Methods Insecticide use, grower preferences regarding genetically engineered (GE) corn resistant to corn rootworm (CRW), and the health effects of using various CRW insecticides (organophosphates, pyrethroids, fipronil and carbamates) are reviewed for current and future farm practices. Results Pest damage to corn has been reduced only one-third by insecticide applications. Health costs from insecticide use appear significant, but costs attributable to CRW control are not quantifiable from available data. Methods reducing health-related costs of insecticide-based CRW control should be evaluated. As a first step, organophosphate insecticide use has been reduced as they have high acute toxicity and risk of long-term neurological consequences. A second step is to use agents which more specifically target the CRW. Conclusion Whereas current insecticides may be poisonous to many species of insects, birds, mammals and humans, a protein derived from Bacillus thurigiensis and produced in plants via genetic modification can target the specific insect of CRW (Coleoptra), sparing other insect and non-insect species from injury.

  20. Exopolysaccharide production by a genetically engineered Enterobacter cloacae strain for microbial enhanced oil recovery.

    Science.gov (United States)

    Sun, Shanshan; Zhang, Zhongzhi; Luo, Yijing; Zhong, Weizhang; Xiao, Meng; Yi, Wenjing; Yu, Li; Fu, Pengcheng

    2011-05-01

    Microbial enhanced oil recovery (MEOR) is a petroleum biotechnology for manipulating function and/or structure of microbial environments existing in oil reservoirs for prolonged exploitation of the largest source of energy. In this study, an Enterobacter cloacae which is capable of producing water-insoluble biopolymers at 37°C and a thermophilic Geobacillus strain were used to construct an engineered strain for exopolysaccharide production at higher temperature. The resultant transformants, GW3-3.0, could produce exopolysaccharide up to 8.83 g l(-1) in molasses medium at 54°C. This elevated temperature was within the same temperature range as that for many oil reservoirs. The transformants had stable genetic phenotype which was genetically fingerprinted by RAPD analysis. Core flooding experiments were carried out to ensure effective controlled profile for the simulation of oil recovery. The results have demonstrated that this approach has a promising application potential in MEOR. Copyright © 2011 Elsevier Ltd. All rights reserved.

  1. Application of micro-genetic algorithm for calibration of kinetic parameters in HCCI engine combustion model

    Institute of Scientific and Technical Information of China (English)

    Haozhong HUANG; Wanhua SU

    2008-01-01

    The micro-genetic algorithm (μGA) as a highly effective optimization method, is applied to calibrate to a newly developed reduced chemical kinetic model (40 species and 62 reactions) for the homogeneous charge compression ignition (HCCI) combustion of n-heptane to improve its autoignition predictions for different engine operating conditions. The seven kinetic parameters of the calibrated model are determined using a combination of the Micro-Genetic Algorithm and the SENKIN program of CHEMKIN chemical kinetics software package. Simulation results show that the autoignition predictions of the calibrated model agree better with those of the detailed chemical kinetic model (544 species and 2 446 reactions) than the original model over the range of equivalence ratios from 0.1-1.3 and temperature from 300-3 000 K. The results of this study have demonstrated that the μGA is an effective tool to facilitate the calibration of a large number of kinetic parameters in a reduced kinetic model.

  2. Mutational breeding and genetic engineering in the development of high grain protein content.

    Science.gov (United States)

    Wenefrida, Ida; Utomo, Herry S; Linscombe, Steve D

    2013-12-04

    Cereals are the most important crops in the world for both human consumption and animal feed. Improving their nutritional values, such as high protein content, will have significant implications, from establishing healthy lifestyles to helping remediate malnutrition problems worldwide. Besides providing a source of carbohydrate, grain is also a natural source of dietary fiber, vitamins, minerals, specific oils, and other disease-fighting phytocompounds. Even though cereal grains contain relatively little protein compared to legume seeds, they provide protein for the nutrition of humans and livestock that is about 3 times that of legumes. Most cereal seeds lack a few essential amino acids; therefore, they have imbalanced amino acid profiles. Lysine (Lys), threonine (Thr), methionine (Met), and tryptophan (Trp) are among the most critical and are a limiting factor in many grain crops for human nutrition. Tremendous research has been put into the efforts to improve these essential amino acids. Development of high protein content can be outlined in four different approaches through manipulating seed protein bodies, modulating certain biosynthetic pathways to overproduce essential and limiting amino acids, increasing nitrogen relocation to the grain through the introduction of transgenes, and exploiting new genetic variance. Various technologies have been employed to improve protein content including conventional and mutational breeding, genetic engineering, marker-assisted selection, and genomic analysis. Each approach involves a combination of these technologies. Advancements in nutrigenomics and nutrigenetics continue to improve public knowledge at a rapid pace on the importance of specific aspects of food nutrition for optimum fitness and health. An understanding of the molecular basis for human health and genetic predisposition to certain diseases through human genomes enables individuals to personalize their nutritional requirements. It is critically important

  3. Recent developments on genetic engineering of microalgae for biofuels and bio-based chemicals.

    Science.gov (United States)

    Ng, I-Son; Tan, Shi-I; Kao, Pei-Hsun; Chang, Yu-Kaung; Chang, Jo-Shu

    2017-08-08

    Microalgae serve as a promising source for the production of biofuels and bio-based chemicals. Microalgae can help mitigate greenhouse effect. They are superior to terrestrial plants as feedstock in many aspects and their biomass is naturally rich in lipids, carbohydrates, proteins, pigments and other valuable compounds. However, there are still some obstacles in developing microalgae-based biofuels and chemicals in industry Due to the relatively slow growth rate and high cultivation cost of microalgae, Therefore, screening of to screen efficient and robust microalgal strains as well as genetic modifications of the available strains for further improvement are of urgent demand in the development of microalgae-based biorefinery. In genetic engineering of microalgae, transformation and selection methods are the key steps to accomplish the target gene modification. For a powerful genetic screening, the resistance gene used should be efficient. However, determination of the preferable type and dosage of antibiotics used for transformant selection is usually time-consuming and microalgal-strain-dependent. Therefore, more powerful and efficient techniques should be developed to meet this need. In this review, the conventional and emerging genome-editing tools (e.g., CRISPR-Cas9, TALEN and ZFN) used in editing the genomes of nuclear, mitochondria and chloroplast of microalgae are thoroughly surveyed. In the current scenario, insufficient genomic data will challenge the applications of such genome editing tools in microalgae. Although all the techniques mentioned above demonstrate their abilities to perform gene editing and desired phenotype screening, there still need to overcome higher production cost and lower biomass productivity, to achieve efficient production of the desired products in microalgal biorefineries. This article is protected by copyright. All rights reserved.

  4. 78 FR 51706 - Bayer CropScience LP; Determination of Nonregulated Status of Soybean Genetically Engineered for...

    Science.gov (United States)

    2013-08-21

    ... regulated article under our regulations governing the introduction of certain genetically engineered....aphis.usda.gov/biotechnology/not_reg.html under APHIS Petition Number 09-328-01p and are posted with the..., Biotechnology Environmental Analysis Branch, Environmental Risk Analysis Programs, Biotechnology Regulatory...

  5. Draft Genome Sequence of the Polycyclic Aromatic Hydrocarbon-Degrading, Genetically Engineered Bioluminescent Bioreporter Pseudomonas fluorescens HK44 ▿

    Science.gov (United States)

    Chauhan, Archana; Layton, Alice C.; Williams, Daniel E.; Smartt, Abby E.; Ripp, Steven; Karpinets, Tatiana V.; Brown, Steven D.; Sayler, Gary S.

    2011-01-01

    Pseudomonas fluorescens strain HK44 (DSM 6700) is a genetically engineered lux-based bioluminescent bioreporter. Here we report the draft genome sequence of strain HK44. Annotation of ∼6.1 Mb of sequence indicates that 30% of the traits are unique and distributed over five genomic islands, a prophage, and two plasmids. PMID:21742869

  6. Draft Genome Sequence of the Polycyclic Aromatic Hydrocarbon-Degrading, Genetically Engineered Bioluminescent Bioreporter Pseudomonas fluorescens HK44

    Energy Technology Data Exchange (ETDEWEB)

    Chauhan, Archana [ORNL; Layton, Alice [University of Tennessee, Knoxville (UTK); Williams, Daniel W [ORNL; Smart, Abby E. [University of Tennessee, Knoxville (UTK); Ripp, Steven Anthony [ORNL; Karpinets, Tatiana V [ORNL; Brown, Steven D [ORNL; Sayler, Gary Steven [ORNL

    2011-01-01

    Pseudomonas fluorescens strain HK44 (DSM 6700) is a genetically engineered lux-based bioluminescent bioreporter. Here we report the draft genome sequence of strain HK44. Annotation of {approx}6.1 Mb sequence indicates that 30% of the traits are unique and distributed over 5 genomic islands, a prophage and two plasmids.

  7. Use of a risk communication model to evaluate dietetics professionals' viewpoints on genetically engineered foods and crops.

    Science.gov (United States)

    Roberts, Kathy S; Struble, Marie Boyle; McCullum-Gomez, Christine; Wilkins, Jennifer L

    2006-05-01

    The complex issues surrounding the application of genetic engineering to food and agriculture have generated a contentious debate among diverse interest groups. One pervasive dimension in the resultant discourse is the varying perceptions of the risks and benefits of genetically engineered foods and crops. In the risk communication model, technical information is evaluated within the context of an individual's values and perceptions. The purpose of this study was to explore how dietetics professionals respond to a complex set of interrelated issues associated with genetically engineered foods and crops and to identify what varying viewpoints may exist. Participants were asked to sort a total of 48 statements distributed across eight issue areas according to level of agreement and disagreement. Using Q methodology, a total of 256 sortings were analyzed using the centroid method and varimax rotation in factor analysis. Three distinct viewpoints emerged: Precautionary (R(2)=43%), Discerning Supporter (R(2)=11%), and Promoting (R(2)=5%). Across all viewpoints, respondents agreed that dietetics professionals should employ critical thinking skills to communicate the social, economic, environmental, ethical, and technical aspects of genetically engineered foods and crops. The findings have implications for how dietetics professionals can foster an open interchange of information among diverse groups.

  8. Draft Genome Sequence of the Polycyclic Aromatic Hydrocarbon-Degrading, Genetically Engineered Bioluminescent Bioreporter Pseudomonas fluorescens HK44 ▿

    OpenAIRE

    Chauhan, Archana; Layton, Alice C.; Williams, Daniel E.; Smartt, Abby E.; Ripp, Steven; Karpinets, Tatiana V.; Brown, Steven D.; Sayler, Gary S.

    2011-01-01

    Pseudomonas fluorescens strain HK44 (DSM 6700) is a genetically engineered lux-based bioluminescent bioreporter. Here we report the draft genome sequence of strain HK44. Annotation of ∼6.1 Mb of sequence indicates that 30% of the traits are unique and distributed over five genomic islands, a prophage, and two plasmids.

  9. 'HoneySweet' (C5), the first genetically engineered Plum pox virus-resistant plum (Prunus domestica L.) cultivar

    Science.gov (United States)

    ‘HoneySweet’ plum was released by the U.S. Department of Agriculture, Agricultural Research Service, to provide U.S. growers and P. domestica plum breeders with a high fruit quality plum cultivar resistant to Plum pox virus (PPV). ‘HoneySweet’ was developed through genetic engineering utilizing the...

  10. Procedures and best management practices for genetically engineered traits in USDA/ARS germplasm and breeding lines

    Science.gov (United States)

    Two decades have passed since the commercialization in the U. S. of crops with genetically engineered (GE) traits. Today more than 80% of corn, soybean, canola, sugar beet and cotton acreage in the United States is planted to transgenic cultivars, but concerns exist regarding how best to manage the ...

  11. Draft genome sequence of the polycyclic aromatic hydrocarbon-degrading, genetically engineered bioluminescent bioreporter Pseudomonas fluorescens HK44.

    Science.gov (United States)

    Chauhan, Archana; Layton, Alice C; Williams, Daniel E; Smartt, Abby E; Ripp, Steven; Karpinets, Tatiana V; Brown, Steven D; Sayler, Gary S

    2011-09-01

    Pseudomonas fluorescens strain HK44 (DSM 6700) is a genetically engineered lux-based bioluminescent bioreporter. Here we report the draft genome sequence of strain HK44. Annotation of ∼6.1 Mb of sequence indicates that 30% of the traits are unique and distributed over five genomic islands, a prophage, and two plasmids.

  12. Engineering control of bacterial cellulose production using a genetic toolkit and a new cellulose-producing strain.

    Science.gov (United States)

    Florea, Michael; Hagemann, Henrik; Santosa, Gabriella; Abbott, James; Micklem, Chris N; Spencer-Milnes, Xenia; de Arroyo Garcia, Laura; Paschou, Despoina; Lazenbatt, Christopher; Kong, Deze; Chughtai, Haroon; Jensen, Kirsten; Freemont, Paul S; Kitney, Richard; Reeve, Benjamin; Ellis, Tom

    2016-06-14

    Bacterial cellulose is a strong and ultrapure form of cellulose produced naturally by several species of the Acetobacteraceae Its high strength, purity, and biocompatibility make it of great interest to materials science; however, precise control of its biosynthesis has remained a challenge for biotechnology. Here we isolate a strain of Komagataeibacter rhaeticus (K. rhaeticus iGEM) that can produce cellulose at high yields, grow in low-nitrogen conditions, and is highly resistant to toxic chemicals. We achieved external control over its bacterial cellulose production through development of a modular genetic toolkit that enables rational reprogramming of the cell. To further its use as an organism for biotechnology, we sequenced its genome and demonstrate genetic circuits that enable functionalization and patterning of heterologous gene expression within the cellulose matrix. This work lays the foundations for using genetic engineering to produce cellulose-based materials, with numerous applications in basic science, materials engineering, and biotechnology.

  13. Application of bone marrow mesenchymal stem cells in cartilage tissue engineering%骨髓间充质干细胞在软骨组织工程中的应用

    Institute of Scientific and Technical Information of China (English)

    黄涛; 吕刚

    2008-01-01

    骨髓基质干细胞应用于软骨组织工程已近20年,这方面的研究也成为干细胞领域的研究热点.本文通过检索文献对骨髓间充质干细胞在软骨组织工程中应用的情况、取得的最新进展及面临的主要问题作一综述.随着分子生物学、生物材料学、计算机及纳米生物技术的发展,以骨髓间充质干细胞为种子细胞复合生物材料支架构建的组织工程软骨在修复关节软骨缺损中将具有广阔的应用前景.%Bone marrow stromal stem cells have been used in cartilage tissue engineering for nearly 20 years. This has been a key focus in stem cell research. This article serves to review application, progress and facing problems of bone marrow mesenchymal stem cells (BMSCs) in cartilage tissue engineering by retrieving publications. With the development of molecular biology, biomaterial, computer and nano-biotechnology, tissue-engineered cartilage constructed with BMSCs as seed cells combined with biomaterial stent has a widely application perspective in repairing articular cartilage defect.

  14. Engineering the Genetic Code in Cells and Animals: Biological Considerations and Impacts.

    Science.gov (United States)

    Wang, Lei

    2017-10-06

    Expansion of the genetic code allows unnatural amino acids (Uaas) to be site-specifically incorporated into proteins in live biological systems, thus enabling novel properties selectively introduced into target proteins in vivo for basic biological studies and for engineering of novel biological functions. Orthogonal components including tRNA and aminoacyl-tRNA synthetase (aaRS) are expressed in live cells to decode a unique codon (often the amber stop codon UAG) as the desired Uaa. Initially developed in E. coli, this methodology has now been expanded in multiple eukaryotic cells and animals. In this Account, we focus on addressing various biological challenges for rewriting the genetic code, describing impacts of code expansion on cell physiology and discussing implications for fundamental studies of code evolution. Specifically, a general method using the type-3 polymerase III promoter was developed to efficiently express prokaryotic tRNAs as orthogonal tRNAs and a transfer strategy was devised to generate Uaa-specific aaRS for use in eukaryotic cells and animals. The aaRSs have been found to be highly amenable for engineering substrate specificity toward Uaas that are structurally far deviating from the native amino acid, dramatically increasing the stereochemical diversity of Uaas accessible. Preparation of the Uaa in ester or dipeptide format markedly increases the bioavailability of Uaas to cells and animals. Nonsense-mediated mRNA decay (NMD), an mRNA surveillance mechanism of eukaryotic cells, degrades mRNA containing a premature stop codon. Inhibition of NMD increases Uaa incorporation efficiency in yeast and Caenorhabditis elegans. In bacteria, release factor one (RF1) competes with the orthogonal tRNA for the amber stop codon to terminate protein translation, leading to low Uaa incorporation efficiency. Contradictory to the paradigm that RF1 is essential, it is discovered that RF1 is actually nonessential in E. coli. Knockout of RF1 dramatically

  15. Visualizing the enteric nervous system using genetically engineered double reporter mice: Comparison with immunofluorescence

    Science.gov (United States)

    Jiang, Yanfen; Dong, Hui; Eckmann, Lars; Hanson, Elaine M.; Ihn, Katherine C.; Mittal, Ravinder K.

    2017-01-01

    Background and aims The enteric nervous system (ENS) plays a crucial role in the control of gastrointestinal motility, secretion and absorption functions. Immunohistochemistry has been widely used to visualize neurons of the ENS for more than two decades. Genetically engineered mice that report specific proteins can also be used to visualize neurons of the ENS. The goal of our study was to develop a mouse that expresses fluorescent neuronal nitric oxide synthase (nNOS) and choline acetyltransferase (ChAT), the two proteins expressed in 95% of the ENS neurons. We compared ENS neurons visualized in the reporter mouse with the wild type mouse stained using classical immunostaining techniques. Methods Mice hemizygous for ChAT-ChR2-YFP BAC transgene with expression of the mhChR2:YFP fusion protein directed by ChAT promoter/enhancer regions on the BAC transgene were purchased commercially. The Cre/LoxP technique of somatic recombination was used to construct mice with nNOS positive neurons. The two mice were crossbred and tissues were harvested and examined using fluorescent microscopy. Immunostaining was performed in the wild type mice, using antibodies to nNOS, ChAT, Hu and PGP 9.5. Results Greater than 95% of the ENS neurons were positive for either nNOS or ChAT or both. The nNOS and ChAT neurons and their processes in the ENS were well visualized in all the regions of the GI tract, i.e., esophagus, small intestine and colon. The number of nNOS and ChAT neurons was approximately same in the reporter mouse and immunostaining method in the wild type mouse. The nNOS fluorescence in the reporter mouse was seen in both cytoplasm as well as nucleus but in the immunostained specimens it was seen only in the cytoplasm. Conclusion We propose that the genetically engineered double reporter mouse for ChAT and nNOS proteins is a powerful tool to study of the effects of various diseases on the ENS without the need for immunostaining. PMID:28158225

  16. Natural and genetically engineered viral agents for oncolysis and gene therapy of human cancers.

    Science.gov (United States)

    Sinkovics, Joseph G; Horvath, Joseph C

    2008-12-01

    Based on personal acquaintances and experience dating back to the early 1950s, the senior author reviews the history of viral therapy of cancer. He points out the difficulties encountered in the treatment of human cancers, as opposed by the highly successful viral therapy of experimentally maintained tumors in laboratory animals, especially that of ascites carcinomas in mice. A detailed account of viral therapy of human tumors with naturally oncolytic viruses follows, emphasizing the first clinical trials with viral oncolysates. The discrepancy between the high success rates, culminating in cures, in the treatment of tumors of laboratory animals, and the moderate results, such as stabilizations of disease, partial responses, very rare complete remissions, and frequent relapses with virally treated human tumors is recognized. The preclinical laboratory testing against established human tumor cell lines that were maintained in tissue cultures for decades, and against human tumors extricated from their natural habitat and grown in xenografts, may not yield valid results predictive of the viral therapy applied against human tumors growing in their natural environment, the human host. Since the recent discovery of the oncosuppressive efficacy of bacteriophages, the colon could be regarded as the battlefield, where incipient tumor cells and bacteriophages vie for dominance. The inner environment of the colon will be the teaching ground providing new knowledge on the value of the anti-tumor efficacy of phage-induced innate anti-tumor immune reactions. Genetically engineered oncolytic viruses are reviewed next. The molecular biology of viral oncolysis is explained in details. Elaborate efforts are presented to elucidate how gene product proteins of oncolytic viruses switch off the oncogenic cascades of cancer cells. The facts strongly support the conclusion that viral therapy of human cancers will remain in the front lines of modern cancer therapeutics. It may be a

  17. A Cell Lysis and Protein Purification - Single Molecule Assay Devices for Evaluation of Genetically Engineered Proteins

    Science.gov (United States)

    Nakyama, Tetsuya; Tabata, Kazuhito; Noji, Hiroyuki; Yokokawa, Ryuji

    We have developed two devices applicable to evaluate genetically engineered proteins in single molecule assay: on-chip cell lysis device, and protein purification - assay device. A motor protein, F1-ATPase expressed in E.coli, was focused in this report as a target protein. Cell lysis was simply performed by applying pulse voltage between Au electrodes patterned by photolithography, and its efficiency was determined by absorptiometry. The subsequent processes, purification and assay of extracted proteins, were demonstrated in order to detect F1-ATPase and to evaluate its activity. The specific bonding between his-tag in F1-ATPase and Ni-NTA coated on a glass surface was utilized for the purification process. After immobilization of F1-ATPase, avidin-coated microspheres and adenosine tri-phosphate (ATP) solution were infused sequentially to assay the protein. Microsphere rotation was realized by activity of F1-ATPase corresponding to ATP hydrolysis. Results show that the cell lysis device, at the optimum condition, extracts enough amount of protein for single molecule assay. Once cell lysate was injected to the purification - assay device, proteins were diffused in the lateral direction in a Y-shape microchannel. The gradient of protein concentratioin provides an optimal concentration for the assay i.e. the highest density of rotating beads. Density of rotating beads is also affected by the initial concentration of protein injected to the device. The optimum concentration was achieved by our cell lysis device not by the conventional method by ultrasonic wave. Rotation speed was analyzed for several microspheres assayed in the purification - assay device, and the results were compatible to that of conventional assay in which F1-ATPase was purified in bulk scale. In conclusion, we have demonstrated on-chip cell lysis and assay appropriate for the sequential analysis without any pretreatment. On-chip devices replacing conventional bioanalytical methods will be

  18. Generation and genetic engineering of human induced pluripotent stem cells using designed zinc finger nucleases.

    Science.gov (United States)

    Ramalingam, Sivaprakash; London, Viktoriya; Kandavelou, Karthikeyan; Cebotaru, Liudmila; Guggino, William; Civin, Curt; Chandrasegaran, Srinivasan

    2013-02-15

    Zinc finger nucleases (ZFNs) have become powerful tools to deliver a targeted double-strand break at a pre-determined chromosomal locus in order to insert an exogenous transgene by homology-directed repair. ZFN-mediated gene targeting was used to generate both single-allele chemokine (C-C motif) receptor 5 (CCR5)-modified human induced pluripotent stem cells (hiPSCs) and biallele CCR5-modified hiPSCs from human lung fibroblasts (IMR90 cells) and human primary cord blood mononuclear cells (CBMNCs) by site-specific insertion of stem cell transcription factor genes flanked by LoxP sites into the endogenous CCR5 locus. The Oct4 and Sox2 reprogramming factors, in combination with valproic acid, induced reprogramming of human lung fibroblasts to form CCR5-modified hiPSCs, while 5 factors, Oct4/Sox2/Klf4/Lin28/Nanog, induced reprogramming of CBMNCs. Subsequent Cre recombinase treatment of the CCR5-modified IMR90 hiPSCs resulted in the removal of the Oct4 and Sox2 transgenes. Further genetic engineering of the single-allele CCR5-modified IMR90 hiPSCs was achieved by site-specific addition of the large CFTR transcription unit to the remaining CCR5 wild-type allele, using CCR5-specific ZFNs and a donor construct containing tdTomato and CFTR transgenes flanked by CCR5 homology arms. CFTR was expressed efficiently from the endogenous CCR5 locus of the CCR5-modified tdTomato/CFTR hiPSCs. These results suggest that it might be feasible to use ZFN-evoked strategies to (1) generate precisely targeted genetically well-defined patient-specific hiPSCs, and (2) then to reshape their function by targeted addition and expression of therapeutic genes from the CCR5 chromosomal locus for autologous cell-based transgene-correction therapy to treat various recessive monogenic human diseases in the future.

  19. Chondrogenic induction of mesenchymal stromal/stem cells from Wharton's jelly embedded in alginate hydrogel and without added growth factor: an alternative stem cell source for cartilage tissue engineering.

    Science.gov (United States)

    Reppel, Loïc; Schiavi, Jessica; Charif, Naceur; Leger, Léonore; Yu, Hao; Pinzano, Astrid; Henrionnet, Christel; Stoltz, Jean-François; Bensoussan, Danièle; Huselstein, Céline

    2015-12-30

    Due to their intrinsic properties, stem cells are promising tools for new developments in tissue engineering and particularly for cartilage tissue regeneration. Although mesenchymal stromal/stem cells from bone marrow (BM-MSC) have long been the most used stem cell source in cartilage tissue engineering, they have certain limits. Thanks to their properties such as low immunogenicity and particularly chondrogenic differentiation potential, mesenchymal stromal/stem cells from Wharton's jelly (WJ-MSC) promise to be an interesting source of MSC for cartilage tissue engineering. In this study, we propose to evaluate chondrogenic potential of WJ-MSC embedded in alginate/hyaluronic acid hydrogel over 28 days. Hydrogels were constructed by the original spraying method. Our main objective was to evaluate chondrogenic differentiation of WJ-MSC on three-dimensional scaffolds, without adding growth factors, at transcript and protein levels. We compared the results to those obtained from standard BM-MSC. After 3 days of culture, WJ-MSC seemed to be adapted to their new three-dimensional environment without any detectable damage. From day 14 and up to 28 days, the proportion of WJ-MSC CD73(+), CD90(+), CD105(+) and CD166(+) decreased significantly compared to monolayer marker expression. Moreover, WJ-MSC and BM-MSC showed different phenotype profiles. After 28 days of scaffold culture, our results showed strong upregulation of cartilage-specific transcript expression. WJ-MSC exhibited greater type II collagen synthesis than BM-MSC at both transcript and protein levels. Furthermore, our work highlighted a relevant result showing that WJ-MSC expressed Runx2 and type X collagen at lower levels than BM-MSC. Once seeded in the hydrogel scaffold, WJ-MSC and BM-MSC have different profiles of chondrogenic differentiation at both the phenotypic level and matrix synthesis. After 4 weeks, WJ-MSC, embedded in a three-dimensional environment, were able to adapt to their environment and

  20. Research Progress on Fish Genetically Engineered Vaccine%鱼用基因工程疫苗研究进展

    Institute of Scientific and Technical Information of China (English)

    田园园; 叶星

    2012-01-01

    疫苗是目前控制鱼类病害最经济有效的方式.免疫学及生物工程的迅速发展极大地促进了鱼类基因工程疫苗的研究.基因工程疫苗克服了传统疫苗的一些缺陷和不足,显示出巨大的应用前景,已成为国内外水产养殖业的研究热点,近年对鱼用基因工程疫苗的研究已取得较大进展,但鱼用基因工程疫苗在研究和应用过程中也面临着急需解决的若干问题.%At present,vaccination is the most cost-effective way to control diseases in fish. The rapid development in immunology and bio-engineering has greatly promoted the studies on genetically engineered vaccines for fish. It overcomes some defects and deficiencies of the traditional vaccines and shows great application prospect,and has become a research focus on aquaculture at home and abroad. This paper reviewed the present status and achievements gained in genetically engineered vaccine against fish pathogens,and problems encountered in commercialization of fishery genetically engineered vaccines that need to be solved urgently.