Synthetic Biology and Engineered Live Biotherapeutics: Toward Increasing System Complexity.

Recent advances in synthetic biology and biological system engineering have allowed the design and construction of engineered live biotherapeutics targeting a range of human clinical applications. In this review, we outline how systems approaches have been used to move from simple constitutive systems, where a single therapeutic molecule is expressed, to systems that incorporate sensing of the in vivo environment, feedback, computation, and biocontainment. We outline examples where each of these capabilities are achieved in different human disorders, including cancer, inflammation, and metabolic disease, in a number of environments, including the gastrointestinal tract, the liver, and the oral cavity. Throughout, we highlight the challenges of developing microbial therapeutics that are both sensitive and specific. Finally, we discuss how these systems are leading to the realization of engineered live biotherapeutics in the clinic.

Engineering Xenopus embryos for phenotypic drug discovery screening.

Many rare human inherited diseases remain untreatable despite the fact that the disease causing genes are known and adequate mouse disease models have been developed. In vivo phenotypic drug screening relies on isolating drug candidates by their ability to produce a desired therapeutic phenotype in whole organisms. Embryos of zebrafish and Xenopus frogs are abundant, small and free-living. They can be easily arrayed in multi-well dishes and treated with small organic molecules. With the development of novel genome modification tools, such a zinc-finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs), and CRISPR/Cas, it is now possible to efficiently engineer non-mammalian models of inherited human diseases. Here, we will review the rapid progress made in adapting these novel genome editing tools to Xenopus. The advantages of Xenopus embryos as in vivo models to study human inherited diseases will be presented and their utility for drug discovery screening will be discussed. Being a tetrapod, Xenopus complements zebrafish as an indispensable non-mammalian animal model for the study of human disease pathologies and the discovery of novel therapeutics for inherited diseases.

ZFN, TALEN, and CRISPR/Cas-based methods for genome engineering.

Zinc-finger nucleases (ZFNs) and transcription activator-like effector nucleases (TALENs) comprise a powerful class of tools that are redefining the boundaries of biological research. These chimeric nucleases are composed of programmable, sequence-specific DNA-binding modules linked to a nonspecific DNA cleavage domain. ZFNs and TALENs enable a broad range of genetic modifications by inducing DNA double-strand breaks that stimulate error-prone nonhomologous end joining or homology-directed repair at specific genomic locations. Here, we review achievements made possible by site-specific nuclease technologies and discuss applications of these reagents for genetic analysis and manipulation. In addition, we highlight the therapeutic potential of ZFNs and TALENs and discuss future prospects for the field, including the emergence of clustered regulatory interspaced short palindromic repeat (CRISPR)/Cas-based RNA-guided DNA endonucleases.

Photothermal genetic engineering.

Optical methods for manipulation of cellular function have enabled deconstruction of genetic and neural circuits in vitro and in vivo. Plasmonic gold nanomaterials provide an alternative platform for external optical manipulation of genetic circuits. The tunable absorption of gold nanoparticles in the infrared spectral region and straightforward surface functionalization has led to applications in intracellular delivery and photorelease of short RNAs, recently enabling bidirectional photothermal modulation of specific genes via RNA interference (RNAi). We discuss recent advances in optical gene circuit engineering and plasmonic nanomaterials, as well as future research opportunities and challenges in photothermal gene manipulation.

Los productos naturales en la búsqueda de nuevos fármacos. Una visión de conjunto / Natural Products as Sources of New Drugs. A general overwiew

Se revisa la importancia de los productos naturales en eldescubrimiento de fármacos y se analizan las ventajas e inconvenientes desu utilización como compuestos modelo. Se discute la idoneidad de estametodología para encontrar fármacos de acción específica, el impacto quepuede originar en los ecosistemas, y los intentos para generar compuestosbioactivos por tramiento de extractos naturales inactivos. Se hace unaespecial mención a los productos naturales de origen marino, se comentanalgunos procesos semisintéticos y se describen ejemplos de los nuevosabordajes que permite la ingeniería genética para optimizar su produccióno lograr la biosíntesis combinatoria(AU)

Natural Products as Sources of New Drugs. A general overwiew.The significance of natural products in drug discovery, as well astheir advantages and drawbacks as model compounds, are examined. Thesuitability of this methodology to find drugs with specific activity, itspossible impact in the ecosystems, and attempts to generate bioactivecompounds through chemically engineered inactive extracts are analyzed.Special mention is given to marine natural products, semisyntheticmethodologies, and description of new approaches to optimize theproduction of natural products or to generate structural diversity throughengineering biosynthetic pathways(AU)

Genetic engineering of radish: current achievements and future goals.

Radish is a major root crop grown in the Far East and is especially important to some low-income countries where it is consumed on a daily basis. Developments in gene technology systems have helped to accelerate the production of useful germplasms, but progress has been slow, though achieved, via in planta methods and useful traits have been introduced. In the wake of the new Millennium, future goals in terms of improving transformation efficiency and selection of new traits for generating late-flowering radish are described. Furthermore, the techniques available for incorporating pharmaceutical proteins into radish to deliver edible proteins on-site are discussed. Finally, the concerns of releasing transgenic radish to the field in terms of pollen-mediated gene transfer are also reviewed. Such a report identifies key areas of research that is required to allow the crop satisfy the need of poor impoverished countries in the Far East.

A genomic approach to nutritional, pharmacological and genetic issues of faba bean (Vicia faba): prospects for genetic modifications.

Cultivated faba bean (Vicia faba) is widely used as human food, especially in Europe, Northern Africa and China.  In view of its superior feeding value over field peas or other legumes, it is also widely used as animal feed for a variety of species.   V. faba also contains medically important components such as 3,4-dihydroxyphenylalanine (levo-DOPA, L-DOPA), the principal treatment used for Parkinson's disease patients.  However, this species also contains several antinutritional components, including the pyrimidine glycosides vicine and convicine; phytates; and the sucrose galactosides including raffinose, stachyose and verbascose.  We have undertaken a genomic project to provide publicly available expressed sequence tag sequences (EST) prepared from early to mid developing embryo in an attempt to identify genes that are likely to be involved in the biosynthesis of L-DOPA and the vicine group of compounds.  As initial examples of the utility of this approach, we describe the complete sequence of fabatin, new defensins, type 4 metallothioneins and a variety of other key genes which were identified in this EST library. No candidate sequences corresponding to the biosynthesis of L-DOPA or the vicine group could be identified at this early stage of seed development.

Biosynthesis and engineering of carotenoids and apocarotenoids in plants: state of the art and future prospects.

Carotenoids and their apocarotenoid derivatives are isoprenoid molecules important for the primary and secondary metabolisms of plants and other living organisms, displaying also key health-related roles in humans and animals. Progress in the knowledge of the carotenoid pathway at the genetic, biochemical and molecular level, supported by successful genetic engineering examples for an increasing number of important plant crops have paved the way for precise molecular breeding of carotenoids. In this review, following a description of the general carotenoid pathway, select examples of plant species able to produce specialty carotenoids and apocarotenoids are illustrated. An update on plant carotenoid engineering is also provided for non-solanaceous crops and members of the Solanaceae family, by means of different strategies and making use of plant and bacterial genes.

Genospirituality: genetic engineering for spiritual and religious enhancement.

The most frequently discussed role for genetic engineering is in relation to medicine, and a second area which provokes discussion is the use of genetic engineering as an enhancement technology. But one neglected area is the potential use of genetic engineering to increase human spiritual and religious experience - or genospirituality. If technologies are devised which can conveniently and safely engineer genes causal of spiritual and religious behaviours, then people may become able to choose their degree of religiosity or spiritual sensitivity. For instance, it may become possible to increase the likelihood of direct religious experience - i.e. 'revelation': the subjective experience of communication from the deity. Or, people may be able to engineer 'animistic' thinking, a mode of cognition in which the significant features of the world - such as large animals, trees, distinctive landscape features - are regarded as sentient and intentional beings; so that the individual experiences a personal relationship with the world. Another potentially popular spiritual ability would probably be shamanism; in which states of altered consciousness (e.g. trances, delirium or dreams) are induced and the shaman may undergo the experience of transformations, 'soul journeys' and contact with a spirit realm. Ideally, shamanistic consciousness could be modulated such that trances were self-induced only when wanted and when it was safe and convenient; and then switched-off again completely when full alertness and concentration are necessary. It seems likely that there will be trade-offs for increased spirituality; such as people becoming less 'driven' to seek status and monetary rewards - as a result of being more spiritually fulfilled people might work less hard and take more leisure. On the other hand, it is also possible that highly moral, altruistic, peaceable and principled behaviours might become more prevalent; and the energy and joyousness of the best churches might spread and be strengthened. Overall, genospirituality would probably be used by people who were unable to have the kind of spiritual or religious experiences which they wanted (or perhaps even needed) in order to lead the kind of life to which they aspired.

Development of agribiotechnology and biosafety regulations used to assess safety of genetically modified crops in Bangladesh.

Bangladesh is on the verge of adopting genetically modified (GM) crops for commercial cultivation and consumption as feed and food. Most of the laboratories are engaged in tissue culture and molecular characterization on plants, whereas some have started living modified organism research with shortages of trained manpower, infrastructure, and funding. Nutritionally improved Golden Rice, biotech brinjal, and late blight-resistant potato are in contained trials in a greenhouse, and potato ring spot virus-resistant papaya is in the process of approval for a field trial. The government has taken some initiative in support of GM organism research, which include the formation of a Biotechnology Department in all institutes and the formation of the apex body, the National Task Force Committee on Biotechnology of Bangladesh under the chairpersonship of the Prime Minister. Biosafety policy guidelines and related aspects of biotechnology issues have been approved, and the laws are in the process of being promulgated. Being a party to the Cartagena Protocol, proper biosafety measures are regulated by the appropriate authority as stated. Although there are no laws made yet directly for biosafety of GM crops/foods, the relevant laws on agriculture, medicine, food, import, trade, environment, etc. may suffice and explain the situation.

Potential applications of human embryonic stem cells.

Advances in reproductive technologies provided opportunity for scientists to be able to grow human embryos in vitro for more than two decades. Skills and knowledge derived from in vitro fertilization and in vitro culture of mammalian embryos opened the chance for scientists to develop the strategies to derive embryonic stem cell lines from mammalian and human embryos. This achievement has initiated a new era in the fields of biotechnology, pharmacology, basic scientific research, and cell-based medicine. To date, scientists have made some progress in optimizing regimens in deriving ES cell lines from human embryos but much more research and development are still required especially in the aspect of directing stem cells into the specific cells of potential clinical use. Collaboration among clinicians and scientists from diverse fields, together with the public awareness of how useful this technology could be to modern medicine, will result in the accumulation of knowledge in this field and, in the near future, a progress in cell-based therapy.

Cooperation of plants and microorganisms: getting closer to the genetic construction of sustainable agro-systems.

The molecular research into two types of beneficial plant-microbe symbioses is reviewed: nutritional (with N(2)-fixing bacteria or mycorrhizal fungi) and defensive (with endo- and epiphytic microbes suppressing pathogens and phytophagans). These symbioses are based on the signaling interactions that result in the development of novel tissue/cellular structures and of extended metabolic capacities in the partners, which greatly improve the adaptive potential of plants due to a decrease in their sensitivity to biotic and abiotic stresses. The molecular, genetic and ecological knowledge on plant-microbe interactions provides a strategy for the organization of sustainable crop production based on substituting the agrochemicals (mineral fertilizers, pesticides) by microbial inoculants. An improvement of plant-microbe symbioses should involve the coordinated modifications in the partners' genotypes resulting in highly complementary combinations. These modifications should be based on the broad utilization of genetic resources from natural symbiotic systems aimed at: (i) increased competitiveness of the introduced (effective) with respect to local (ineffective) microbial strains, and (ii) overcoming the limiting steps in the metabolic machineries of the symbiotic systems.

Genetic engineering and functional foods / Engenharia genética e alimentos funcionais

Functional foods are the food-industry response to the continuous ly increasing request of consumers for foods that are both attractive and healthy. The main targets of functional foods are intestinal health, immune system activity, mental performance, caries, menopause symptoms, cancer, cardiovascular disease, diabetes, osteoporosis and child skeletal development. Most of the functional foods designed so far are derived from traditional foods by adding so-called functional ingredients, by modifying the technological process during industrial food preparation or by modifying the composition of the raw material used for food production. However, gene technology is thought to be a powerful technique to improve the nutritional quality of food raw materials. The modification of product quality characteristics using gene technology depends on a well-establishe dunder standing of the pathways for biosynthesis of plant products, a rapidly expanding knowledge about the genetic control of these pathways, and an increasing availability of cloned genes for key enzymatic steps. Quality-improved crops derived from genetic engineering are expected to reach the market in the near future. Crops with an improved protein quality, with an improved nutritional quality of the plant oil, crops rich in vitamins, minerals, antioxidants or low in undesired compounds as well as crops with an altered secondary metabolite production or altered carbohydrate composition have been developed by genetic engineering. These examples give an idea of the genetic engineering potential to produce health-promoting foods

Los alimentos funcionales son la respuesta de la industria de alimentos a la creciente demanda de los consumidores por alimentos que sean al mismo tiempo atrayentes y saludables. Los principales objetivos de los alimentos funcionales son la salud intestinal, del sistema inmunológico,el desempeño mental, las caries, los síntomas dela menopausia, cáncer, enfermidades cardiovasculares, diabetes, osteoporosis y desenvolvimiento óseo en niños. La mayoría de los alimentos funcionales desarrollados hasta el momento son derivados de los alimentos tradicionales a los cuales se les adicionan los ingredientes funcionales, se les modifica el proceso tecnológico de industrialización o se les altera la composición de materias primas utilizadas en su producción. Sin embargo, se tiene por cierto que la tecnología genética es un instrumento poderoso para mejorar la calidad nutricional de las materias primas alimenticias. La modificación de las características de calidad del producto utilizando tecnología genética depende de un conocimiento asentado de las vías metabólicas de síntesis de productos vegetales, un conocimiento en rápida expansión sobre el control genético de tales vías y una creciente disponibilidad de genes clonados para la expresión de enzimas claves de algunos passos de esas vías. Se espera que cultivos con calidad mejorada originarios de ingeniería genéticalleguen al mercado en un futuro próximo.Cultivos con mejor calidad de proteínas y lípidos, con mayor concentración de vitaminas, minerales y antioxidantes, con bajos tenores decompuestos indeseables y también cultivos com metabolitos secundarios modificados o composición alterada de carbohidratos son ejemplos de logros ya alcanzados por la ingeniería genética. Los ejemplos mencionados permiten visualizar el potencial de la ingeniería genética para la producción de alimentos promotores de la salud

Os alimentos funcionais são a resposta da indústria alimentícia à sempre crescente demanda dos consumidores por alimentos ao mesmo tempo atraentes e saudáveis. Os principais alvos dos alimentos funcionais são a saúde intestinal, a atividade do sistema imune, o desempenho mental, cáries, sintomas da menopausa, câncer, doenças cardiovasculares, diabetes, osteoporose e desenvolvimento ósseo de crianças. A maioria dos alimentos funcionais desenvolvidos, até o momento, são derivados de alimentos tradicionais pela adição dos ditos ingredientes funcionais, modificação dos processos tecnológicos durante o preparo industrial dos alimentos ou alteração da composição das matérias-primas usadas na produção dos alimentos. Contudo, acredita-se que a tecnologia genética seja um poderoso instrumento para melhorar a qualidade nutricional das matérias-primas alimentícias. A modificação das características de qualidade do produto usando a tecnologia genética depende de um conhecimento bem embasado sobre as rotas metabólicas de síntese de produtos vegetais, um conhecimento em rápida expansão sobre o controle genético de tais rotas metabólicas, e uma crescente disponibilidade de genes clonados para expressão de enzimas-chave de alguns passos destas rotas. Espera-se que culturas com qualidade melhorada derivadas da engenharia genética cheguem ao mercado num futuro próximo. Culturas com qualidade proteica melhorada, com melhor qualidade nutricional do óleo vegetal derivado, culturas ricas em vitaminas, minerais, antioxidantes ou com baixos teores de compostos indesejáveis, bem como culturas com produção de metabólitos secundários alterados ou composição alterada de carboidratos já foram desenvolvidas pela engenharia genética. Estes exemplos dão uma ideia do potencial da engenharia genética para produzir alimentos promotores de saúde

Plant protoplasts: status and biotechnological perspectives.

Plant protoplasts ("naked" cells) provide a unique single cell system to underpin several aspects of modern biotechnology. Major advances in genomics, proteomics, and metabolomics have stimulated renewed interest in these osmotically fragile wall-less cells. Reliable procedures are available to isolate and culture protoplasts from a range of plants, including both monocotyledonous and dicotyledonous crops. Several parameters, particularly the source tissue, culture medium, and environmental factors, influence the ability of protoplasts and protoplast-derived cells to express their totipotency and to develop into fertile plants. Importantly, novel approaches to maximise the efficiency of protoplast-to-plant systems include techniques already well established for animal and microbial cells, such as electrostimulation and exposure of protoplasts to surfactants and respiratory gas carriers, especially perfluorochemicals and hemoglobin. However, despite at least four decades of concerted effort and technology transfer between laboratories worldwide, many species still remain recalcitrant in culture. Nevertheless, isolated protoplasts are unique to a range of experimental procedures. In the context of plant genetic manipulation, somatic hybridisation by protoplast fusion enables nuclear and cytoplasmic genomes to be combined, fully or partially, at the interspecific and intergeneric levels to circumvent naturally occurring sexual incompatibility barriers. Uptake of isolated DNA into protoplasts provides the basis for transient and stable nuclear transformation, and also organelle transformation to generate transplastomic plants. Isolated protoplasts are also exploited in numerous miscellaneous studies involving membrane function, cell structure, synthesis of pharmaceutical products, and toxicological assessments. This review focuses upon the most recent developments in protoplast-based technologies.

Lessons we can learn from ecological biosafety research.

The last decade has seen an increasing number of biosafety related publications focusing on transgenic organisms. Recent extensive field studies suggest that harmful laboratory effects on non-target organisms rarely occur in the environment. Moreover, biosafety studies typically show no difference in hybridisation between genetically modified plants (GMPs) or non-GMPs and related wild species. Since risk is a product of both exposure and hazard, biosafety research should clearly not only target gene flow exposure but specifically concentrate on expected hazards emerging from successful transgene flow to wild relatives of GMPs. Generally, transgenic plants behave in an ecologically similar manner to non-GMPs if the modified trait confers a neutral advantage under environmental or experimental conditions. However, GMPs perform better than non-GMPs if the new phenotype is challenged by conditions ecologically advantageous for the modified trait. Since biosafety research is a laborious process it will have to concentrate resources on thoughtful, thorough experiments, and target ecologically 'riskier' organisms. So far, we have no evidence that the use of GMPs contradicts sustainable agriculture and nature conservation per se.

Genetic technologies and achieving health for populations.

The remarkable progress in genetics over the last 50 years has led to the development of genetic technologies to identify or alter genes in living organisms, and these technologies can be applied to people. This article presents background information on the role of genetics in human disease, outlines the technologies, and discusses the sources of the strong push for a genetic approach to ill-health and some implications and harmful consequences of using these genetic technologies. The determinants of most diseases are complex and are embedded in a social context. To focus on only one strand of this web--the genetic strand--because it is one that may be amenable to biological/pharmaceutical treatment, although profitable for industry, does not address other important determinants of health and may lead to a harmful overemphasis on genetic approaches. The author outlines some limitations to the potential contribution of genetic technologies to population health across the globe and the need for policy development if these technologies are to have an appropriate place in health care.

Applications of the green fluorescent protein in cell biology and biotechnology.

The recent emergence of an autofluorescent protein, the green fluorescent protein (GFP), has opened the door for the convenient use of intact living cells and organisms as experimental systems in fields ranging from cell biology to biomedicine. We present an overview of some of the major applications of GFP, namely its use in protein tagging and in monitoring gene expression as well as its potential in a variety of biological screens.

Future prospects for genetic modification of the composition of edible oils from higher plants.

It is now routinely possible to introduce genes into many plant species of agronomic significance. This has created new opportunities to genetically engineer higher plants to produce edible fats and oils with predefined fatty acid composition. Because of the chemical diversity of plants, the genes required for synthesis of many different types of lipids exist in nondomesticated species. Thus, it should be possible to modify the storage-lipid composition of crop plants by transferring the relevant genes from the wild species into crop plants. However, although a coherent model now exists for plant-lipid metabolism, a substantial amount of the specific information required to undertake genetic engineering of plant-lipid metabolism is not yet available.