Enhanced Genetic Tools for Engineering Multigene Traits into Green Algae

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.     

The past,present and future of phycology in China

Algae have been part of Chinese life for thousands of years. They are widely used as food and have been cited in Chinese literature as early as 2500 years ago. However, formal taxonomic studies on Chinese algae were initiated by foreign scientists only about 200 years ago, and by Chinese phycologists only about 90 years ago. This paper summarizes the history of modern phycological studies on Chinese algae and provides an overview of the achievements of phycological studies by Chinese scientists, especially on algal taxonomy, morphology, genetics, ecology and environmental research, physiology, biotechnology, algal culture, applied phycology and space phycology, in the last century. Recent development in phycological research focuses on algal floristic and molecular systematics, algal molecular biotechnology, applied phycology including micro and macroalgal cultivation and algal product development, and the roles of algae in environmental pollution control. These areas will also be the main focuses of Chinese phycological research in the foreseeable future.     

COMMERCIAL DEVELOPMENTS IN MICROALGAL BIOTECHNOLOGY

A number of important advances have occurred in microalgal biotechnology in recent years that are slowly moving the field into new areas. New products are being developed for use in the mass commercial markets as opposed to the "health food" markets. These include algal-derived long-chained polyunsaturated fatty acids, mainly docosahexaenoic acid, for use as supplements in human nutrition and animals. Large-scale production of algal fatty acids is possible through the use of heterotrophic algae and the adaptation of classical fermentation systems providing consistent biomass under highly controlled conditions that result in a very high quality product. New products have also been developed for use in the development of pharmaceutical and research products. These include stable-isotope biochemicals produced by algae in closed-system photobioreactors and extremely bright fluorescent pigments. Cryopreservation has also had a tremendous impact on the ability of strains to be maintained for long periods of time at low cost and maintenance while preserving genetic stability.     

Recent patents in plant biotechnology: impact on global health

Agricultural biotechnology offers a robust series of tools by which to address global concerns such as food security, crop protection, and fuel/energy requirements. A number of advances made recently in plant molecular biology also have resulted in applications which largely focus on improving global human health. This review describes some of the recent innovations in plant biotechnology that have come to the forefront over the past year. Included are novel techniques by which plants can be improved as platforms for biopharmaceutical protein production, a growing field also referred to as 'molecular pharming'. The metabolic engineering of plants to produce compounds which have additional nutritional benefits is also outlined. The review concludes with a discussion of the future impact that these innovations may have both on global health and on the development of our future intellectual property landscape.     

低能离子束生物技术的应用

自从我国科学家发现离子注入生物学效应后,低能离子束生物技术的研究就在我国率先兴起。随后,越来越多的科学家基于低能离子与生物体之间存在的能量沉积、动量传递、质量沉积及电荷中和与交换的相互作用,对生物体内的遗传物质进行加工、修饰、重组,开辟了农作物和微生物等遗传改良及转基因的新方法。本文简要介绍了低能离子束生物技术产生的背景、低能离子束与生物体之间相互作用的机理和特点以及目前低能离子束在诱变育种和转基因等生物技术领域的研究进展,并展望了离子束技术在藻类基因工程方面的发展潜力。     

Opportunities for yeast metabolic engineering: Lessons from synthetic biology

Constant progress in genetic engineering has given rise to a number of promising areas of research that facilitated the expansion of industrial biotechnology. The field of metabolic engineering, which utilizes genetic tools to manipulate microbial metabolism to enhance the production of compounds of interest, has had a particularly strong impact by providing new platforms for chemical production. Recent developments in synthetic biology promise to expand the metabolic engineering toolbox further by creating novel biological components for pathway design. The present review addresses some of the recent advances in synthetic biology and how these have the potential to affect metabolic engineering in the yeast Saccharomyces cerevisiae. While S. cerevisiae for years has been a robust industrial organism and the target of multiple metabolic engineering trials, its potential for synthetic biology has remained relatively unexplored and further research in this field could strongly contribute to industrial biotechnology. This review also addresses are general considerations for pathway design, ranging from individual components to regulatory systems, overall pathway considerations and whole-organism engineering, with an emphasis on potential contributions of synthetic biology to these areas. Some examples of applications for yeast synthetic biology and metabolic engineering are also discussed.     

Road to the future of systems biotechnology: CRISPR-Cas-mediated metabolic engineering for recombinant protein production

The rising potential for CRISPR–Cas-mediated genome editing has revolutionized our strategies in basic and practical bioengineering research. It provides a predictable and precise method for genome modification in a robust and reproducible fashion. Emergence of systems biotechnology and synthetic biology approaches coupled with CRISPR–Cas technology could change the future of cell factories to possess some new features which have not been found naturally. We have discussed the possibility and versatile potentials of CRISPR–Cas technology for metabolic engineering of a recombinant host for heterologous protein production. We describe the mechanisms involved in this metabolic engineering approach and present the diverse features of its application in biotechnology and protein production.     

Biochemistry,genetics and biotechnology of glycerol utilization in Pseudomonas species

The use of renewable waste feedstocks is an environment-friendly choice contributing to the reduction of waste treatment costs and increasing the economic value of industrial by-products. Glycerol (1,2,3-propanetriol), a simple polyol compound widely distributed in biological systems, constitutes a prime example of a relatively cheap and readily available substrate to be used in bioprocesses. Extensively exploited as an ingredient in the food and pharmaceutical industries, glycerol is also the main by-product of biodiesel production, which has resulted in a progressive drop in substrate price over the years. Consequently, glycerol has become an attractive substrate in biotechnology, and several chemical commodities currently produced from petroleum have been shown to be obtained from this polyol using whole-cell biocatalysts with both wild-type and engineered bacterial strains. Pseudomonas species, endowed with a versatile and rich metabolism, have been adopted for the conversion of glycerol into value-added products (ranging from simple molecules to structurally complex biopolymers, e.g. polyhydroxyalkanoates), and a number of metabolic engineering strategies have been deployed to increase the number of applications of glycerol as a cost-effective substrate. The unique genetic and metabolic features of glycerol-grown Pseudomonas are presented in this review, along with relevant examples of bioprocesses based on this substrate – and the synthetic biology and metabolic engineering strategies implemented in bacteria of this genus aimed at glycerol valorization.     

Industrial biotechnology: Tools and applications

Industrial biotechnology involves the use of enzymes and microorganisms to produce value-added chemicals from renewable sources. Because of its association with reduced energy consumption, greenhouse gas emissions, and waste generation, industrial biotechnology is a rapidly growing field. Here we highlight a variety of important tools for industrial biotechnology, including protein engineering, metabolic engineering, synthetic biology, systems biology, and downstream processing. In addition, we show how these tools have been successfully applied in several case studies, including the production of 1, 3-propanediol, lactic acid, and biofuels. It is expected that industrial biotechnology will be increasingly adopted by chemical, pharmaceutical, food, and agricultural industries.     

ALGAL TRANSGENICS IN THE GENOMIC ERA1

The last few years have witnessed significant advances in the field of algal genomics. Complete genome sequences from the red alga Cyanidioschyzon merolae and the diatom Thalassiosira pseudonana have been published, the genomes for two more algae (Chlamydomonas reinhardtii and Ostreococcus tauri) are nearing completion, and several others are in progress or at the planning stage. In addition, large‐scale cDNA sequencing projects are being carried out for numerous algal species. This wealth of genome data is serving as a powerful catalyst for the development and application of recombinant techniques for these species. The data provide a rich resource of DNA elements such as promoters that can be used for transgene expression as well as an inventory of genes that are possible targets for genetic engineering programs aimed at manipulating algal metabolism. It is not surprising therefore that significant progress in the genetic engineering of eukaryotic algae is being made. Nuclear transformation of various microalgal species is now routine, and progress is being made on the transformation of macroalgae. Chloroplast transformation has been achieved for green, red, and euglenoid algae, and further success in organelle transformation is likely as the number of sequenced plastid, mitochondrial, and nucleomorph genomes continues to grow. Importantly, the commercial application of algal transgenics is beginning to be realized, and algal biotechnology companies are being established. Recent work has shown that recombinant proteins of therapeutic value can be produced in microalgal species, and it is now realistic to envisage the genetic engineering of commercially important species to improve production of valuable algal products. In this article we review the recent progress in algal transgenics and consider possible future developments now that phycology has entered the genomic era.     

Aspergillus as a multi-purpose cell factory: current status and perspectives

Aspergilli have a long history in biotechnology as expression platforms for the production of food ingredients, pharmaceuticals and enzymes. The achievements made during the last years, however, have the potential to revolutionize Aspergillus biotechnology and to assure Aspergillus a dominant place among microbial cell factories. This mini-review will highlight most recent breakthroughs in fundamental and applied Aspergillus research with a focus on new molecular tools, techniques and products. New trends and concepts related to Aspergillus genomics and systems biology will be discussed as well as the challenges that have to be met to integrate omics data with metabolic engineering attempts.     

Development and enhancement of agricultural biotechnology in some countries in Latin America

A number of research institutions and both local and international agencles in Latin America are using biotechnology as part of an effort to enhance agricultural productivity. However, it is very much an open question as to whether all of these various organizations can provide the best means of realizing this goal. Latin American countries vary dramatically in their knowledge base and current use of modern biotechnology. Thus, while some countries lack the ability to develop, or possibly even implement, many aspects of modern biotechnology, others are quite advanced in this regard. This review provides a somewhat selective overview of current research in the area of agricultural biotechnology in Mexico, Costa Rica and Ecuador, with emphasis on how the existing agencies and institutions have responded to the challenge of biotechnology. In addition, general strategies for the development of agricultural biotechnology in these countries are presented and discussed.     

专家论藻类学的新前沿

随着可供利用的陆生资源的日益枯竭以及系统生物学与整合生物技术的兴起,藻类学研究正面临着许多前所未有的挑战与机遇。在312次香山科学会议——《藻类学的新前沿》上,与会专家围绕“系统生物学时代的藻类学研究”、“面向资源环境新需求的藻类生物技术”、“藻类学科和藻类产业的可持续发展”、“藻类与环境”等关键问题,进行了深入的探讨。     

The evolution of the biotechnology industry in Germany

In the past five years, the climate for commercial biotechnology in Germany has improved significantly and has resulted in an increase in the number of biotechnology companies. On examination of the underlying factors of the evolution of the biotechnology industry in Germany, and against the background of the current situation, it is predicted that many German biotech companies will have to change their business models to focus on product development rather than on platform technologies.     

Striking convergence in the mouthpart evolution of stream-living algae grazers

This scanning-electron microscopic study demonstrates the convergent evolution of the mouthparts of various herbivorous stream animals (insects from different orders, an isopod, snails, fish, and a tadpole) feeding on epilithic algal pastures. This food source is rich but is often difficult to harvest. Nevertheless, a large number of species can live on it because they have evolved highly specialized mouthparts. There are four functional problems that an algae grazer has to solve: the algae must be removed from the stone, they have to be collected and crushed, and a current shield is needed to prevent the water flow sweeping away the food. Among the 30 algae grazers examined in this study, a limited number of morphological solutions have been found for each of these adaptational problems. There are multiple evolutionary pathways for mouthpart adaptation and even closely related species have often evolved different types of tools for the same function. This refects the existence of a certain amount of evolutionary scope. Such freedom of evolution is present, however, only at the beginning of the adaptiogenesis of an algae grazer. Once one of the evolutionary pathways is taken, further improvement of the mouthparts is possible only by the refinement of the ‘chosen’ type of tools. The consequence of this is that a large number of astonishing convergences have occurred in algae grazers that have independently trodden the same evolutionary pathway.     

Bio-mining the microbial treasures of the ocean: new natural products

The biological resources of the oceans have been exploited since ancient human history, mainly by catching fish and harvesting algae. Research on natural products with special emphasis on marine animals and also algae during the last decades of the 20th century has revealed the importance of marine organisms as producers of substances useful for the treatment of human diseases. Though a large number of bioactive substances have been identified, some many years ago, only recently the first drugs from the oceans were approved. Quite astonishingly, the immense diversity of microbes in the marine environments and their almost untouched capacity to produce natural products and therefore the importance of microbes for marine biotechnology was realized on a broad basis by the scientific communities only recently. This has strengthened worldwide research activities dealing with the exploration of marine microorganisms for biotechnological applications, which comprise the production of bioactive compounds for pharmaceutical use, as well as the development of other valuable products, such as enzymes, nutraceuticals and cosmetics. While the focus in these fields was mainly on marine bacteria, also marine fungi now receive growing attention. Although culture-dependent studies continue to provide interesting new chemical structures with biological activities at a high rate and represent highly promising approaches for the search of new drugs, exploration and use of genomic and metagenomic resources are considered to further increase this potential. Many efforts are made for the sustainable exploration of marine microbial resources. Large culture collections specifically of marine bacteria and marine fungi are available. Compound libraries of marine natural products, even of highly purified substances, were established. The expectations into the commercial exploitation of marine microbial resources has given rise to numerous institutions worldwide, basic research facilities as well as companies. In Europe, recent activities have initiated a dynamic development in marine biotechnology, though concentrated efforts on marine natural product research are rare. One of these activities is represented by the Kieler Wirkstoff-Zentrum KiWiZ, which was founded in 2005 in Kiel (Germany).     

Recombinant viruses as transformation vectors of marine macroalgae

The large dsDNA viruses that are known to infect eukaryotic algae show promise as genetic vectors for algal biotechnology. The large size (150–330 kbp) of these viral genomes may permit insertion of large sequences of foreign DNA. The viruses infecting filamentous marine brown algae appear to be integrated into the genomes of their hosts, and may provide integration mechanisms that can be used for directing insertion of foreign genes into algal chromosomes.     

Biodiesel-fuel: Content,production, producers,contemporary biotechnology (Review)

The necessity of expanding studies on producing renewable biofuel is reviewed. Special attention is given to biodiesel, the history of its creation, and its advantages and disadvantages in comparison with diesel-fuel. The main part of the review is devoted to an analysis of diesel biofuel on the basis of bacterial lipids, filamentous fungi, yeasts, plants, photo- and heterotrophic algae. Biodiesel on the basis of filamentous fungi is studied in detail and the possibility of creation of the most perspective biotechnology using these producers is grounded. The contemporary state of biotechnology in Russia is discussed in connection with the development of energetics based on renewable biofuels.     

The genome‐scale metabolic network of Ectocarpus siliculosus (EctoGEM): a resource to study brown algal physiology and beyond

Brown algae (stramenopiles) are key players in intertidal ecosystems, and represent a source of biomass with several industrial applications. Ectocarpus siliculosus is a model to study the biology of these organisms. Its genome has been sequenced and a number of post‐genomic tools have been implemented. Based on this knowledge, we report the reconstruction and analysis of a genome‐scale metabolic network for E. siliculosus, EctoGEM ( http://ectogem.irisa.fr ). This atlas of metabolic pathways consists of 1866 reactions and 2020 metabolites, and its construction was performed by means of an integrative computational approach for identifying metabolic pathways, gap filling and manual refinement. The capability of the network to produce biomass was validated by flux balance analysis. EctoGEM enabled the reannotation of 56 genes within the E. siliculosus genome, and shed light on the evolution of metabolic processes. For example, E. siliculosus has the potential to produce phenylalanine and tyrosine from prephenate and arogenate, but does not possess a phenylalanine hydroxylase, as is found in other stramenopiles. It also possesses the complete eukaryote molybdenum co‐factor biosynthesis pathway, as well as a second molybdopterin synthase that was most likely acquired via horizontal gene transfer from cyanobacteria by a common ancestor of stramenopiles. EctoGEM represents an evolving community resource to gain deeper understanding of the biology of brown algae and the diversification of physiological processes. The integrative computational method applied for its reconstruction will be valuable to set up similar approaches for other organisms distant from biological benchmark models.