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1.
Novel biotechnological approaches to produce biological compounds: challenges and opportunities for science communication.
Pei, L, Schmidt, M
Current opinion in biotechnology. 2019;:43-47
Abstract
Novel biotechnological approaches such as Metabolic Engineering (ME) and New Plant Breeding Techniques (NPBTs) are currently being developed to produce biological compounds for food and non-food products. NPBTs span a range of methods for in vivo production in crops, some of which are classified as GMOs while others aren't. Deploying such techniques will not only provide new opportunities for industry, but also challenges with respect to the regulatory environment. Similarly, the process of communicating these new techniques and their products to stakeholders and consumers will not be without its own challenges. We argue that scientists should engage more with non-scientists, either directly or through collaborators. These engagements should not only be about the science, we suggest, but also explicitly deal with real world ramifications, such as economic, environmental and social issues.
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2.
Engineering Translation Components Improve Incorporation of Exotic Amino Acids.
Katoh, T, Suga, H
International journal of molecular sciences. 2019;(3)
Abstract
Methods of genetic code manipulation, such as nonsense codon suppression and genetic code reprogramming, have enabled the incorporation of various nonproteinogenic amino acids into the peptide nascent chain. However, the incorporation efficiency of such amino acids largely varies depending on their structural characteristics. For instance, l-α-amino acids with artificial, bulky side chains are poorer substrates for ribosomal incorporation into the nascent peptide chain, mainly owing to the lower affinity of their aminoacyl-tRNA toward elongation factor-thermo unstable (EF-Tu). Phosphorylated Ser and Tyr are also poorer substrates for the same reason; engineering EF-Tu has turned out to be effective in improving their incorporation efficiencies. On the other hand, exotic amino acids such as d-amino acids and β-amino acids are even poorer substrates owing to their low affinity to EF-Tu and poor compatibility to the ribosome active site. Moreover, their consecutive incorporation is extremely difficult. To solve these problems, the engineering of ribosomes and tRNAs has been executed, leading to successful but limited improvement of their incorporation efficiency. In this review, we comprehensively summarize recent attempts to engineer the translation systems, resulting in a significant improvement of the incorporation of exotic amino acids.
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3.
Cell-free gene-regulatory network engineering with synthetic transcription factors.
Swank, Z, Laohakunakorn, N, Maerkl, SJ
Proceedings of the National Academy of Sciences of the United States of America. 2019;(13):5892-5901
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Abstract
Gene-regulatory networks are ubiquitous in nature and critical for bottom-up engineering of synthetic networks. Transcriptional repression is a fundamental function that can be tuned at the level of DNA, protein, and cooperative protein-protein interactions, necessitating high-throughput experimental approaches for in-depth characterization. Here, we used a cell-free system in combination with a high-throughput microfluidic device to comprehensively study the different tuning mechanisms of a synthetic zinc-finger repressor library, whose affinity and cooperativity can be rationally engineered. The device is integrated into a comprehensive workflow that includes determination of transcription-factor binding-energy landscapes and mechanistic modeling, enabling us to generate a library of well-characterized synthetic transcription factors and corresponding promoters, which we then used to build gene-regulatory networks de novo. The well-characterized synthetic parts and insights gained should be useful for rationally engineering gene-regulatory networks and for studying the biophysics of transcriptional regulation.
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4.
Harnessing CRISPR-Cas systems for precision engineering of designer probiotic lactobacilli.
Goh, YJ, Barrangou, R
Current opinion in biotechnology. 2019;:163-171
Abstract
Our evolving understanding on the mechanisms underlying the health-promoting attributes of probiotic lactobacilli, together with an expanding genome editing toolbox have made this genus an ideal chassis for the development of living therapeutics. The rising adoption of CRISPR-based technologies for prokaryotic engineering has demonstrated precise, efficient and scalable genome editing and tunable transcriptional regulation that can be translated into next-generation development of probiotic lactobacilli with enhanced robustness and designer functionalities. Here, we discuss how these tools in conjunction with the naturally abundant and diverse native CRISPR-Cas systems can be harnessed for Lactobacillus cell surface engineering and the delivery of biotherapeutics.
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Report on the SCRA Nuts and Bolts Workshop II: case studies of citrus greening, Ultra-low Gossypol Cotton, and blight tolerant, low-acrylamide potato.
Hood, EE, Eversole, KA, Leach, L, Hogan, M, McHughen, A, Cordts, J, Rathore, K, Rood, T, Collinge, S, Irey, M
GM crops & food. 2019;(3):139-158
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Abstract
To be commercialized and grown in the US, genetically engineered (GE) crops typically go through an extensive food, feed, and environmental safety assessment process which, in certain instances, requires complex consultations with three different US regulatory agencies. Many small market, niche, and specialty crops have been genetically engineered using the modern tools of recombinant DNA but few have been commercialized due to real or perceived regulatory constraints. This workshop discussed the practical aspects of developing dossiers on GE specialty, niche, or small-market crops/products for submission to US regulatory agencies. This workshop focused on actual case studies, and provided an opportunity for public or private sector scientists and crop developers to spend time with regulatory officials to learn the specifics of compiling a dossier for regulatory approval. The objective of the workshop was to explain and demystify data requirements and regulatory dossier compilation by small companies, academics, and other developers.
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6.
Genetic Engineering for Disease Resistance in Plants: Recent Progress and Future Perspectives.
Dong, OX, Ronald, PC
Plant physiology. 2019;(1):26-38
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Abstract
A review of the recent progress in plant genetic engineering for disease resistance highlights future challenges and opportunities in the field.
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7.
Breeding crops to feed 10 billion.
Hickey, LT, N Hafeez, A, Robinson, H, Jackson, SA, Leal-Bertioli, SCM, Tester, M, Gao, C, Godwin, ID, Hayes, BJ, Wulff, BBH
Nature biotechnology. 2019;(7):744-754
Abstract
Crop improvements can help us to meet the challenge of feeding a population of 10 billion, but can we breed better varieties fast enough? Technologies such as genotyping, marker-assisted selection, high-throughput phenotyping, genome editing, genomic selection and de novo domestication could be galvanized by using speed breeding to enable plant breeders to keep pace with a changing environment and ever-increasing human population.
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8.
Capsid Engineering Overcomes Barriers Toward Adeno-Associated Virus Vector-Mediated Transduction of Endothelial Cells.
Zhang, L, Rossi, A, Lange, L, Meumann, N, Koitzsch, U, Christie, K, Nesbit, MA, Moore, CBT, Hacker, UT, Morgan, M, et al
Human gene therapy. 2019;(10):1284-1296
Abstract
Endothelial cells (EC) are targets in gene therapy and regenerative medicine, but they are inefficiently transduced with adeno-associated virus (AAV) vectors of various serotypes. To identify barriers hampering efficient transduction and to develop an optimized AAV variant for EC transduction, we screened an AAV serotype 2-based peptide display library on primary human macrovascular EC. Using a new high-throughput selection and monitoring protocol, we identified a capsid variant, AAV-VEC, which outperformed the parental serotype as well as first-generation targeting vectors in EC transduction. AAV vector uptake was improved, resulting in significantly higher transgene expression levels from single-stranded vector genomes detectable within a few hours post-transduction. Notably, AAV-VEC transduced not only proliferating EC but also quiescent EC, although higher particle-per-cell ratios had to be applied. Also, induced pluripotent stem cell-derived endothelial progenitor cells, a novel tool in regenerative medicine and gene therapy, were highly susceptible toward AAV-VEC transduction. Thus, overcoming barriers by capsid engineering significantly expands the AAV tool kit for a wide range of applications targeting EC.
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The application of the CRISPR-Cas9 genome editing machinery in food and agricultural science: Current status, future perspectives, and associated challenges.
Eş, I, Gavahian, M, Marti-Quijal, FJ, Lorenzo, JM, Mousavi Khaneghah, A, Tsatsanis, C, Kampranis, SC, Barba, FJ
Biotechnology advances. 2019;(3):410-421
Abstract
The recent progress in genetic engineering has brought multiple benefits to the food and agricultural industry by enhancing the essential characteristics of agronomic traits. Powerful tools in the field of genome editing, such as siRNA-mediated RNA interference for targeted suppression of gene expression and transcription activator-like effector nucleases (TALENs) and zinc-finger nucleases (ZFNs) for DNA repair have been widely used for commercial purposes. However, in the last few years, the discovery of the CRISPR-Cas9 system has revolutionized genome editing and has attracted attention as a powerful tool for several industrial applications. Herein, we review current progresses in the utilization of the CRISPR-Cas9 system in the food and agricultural industry, particularly in the development of resistant crops with improved quality and productivity. We compare the CRISPR system with the TALEN and ZFN nucleases-based methods and highlight potential advantages and shortcomings. In addition, we explore the state of the global market and discuss the safety and ethical concerns associated with the application of this technology in the food and agricultural industry.
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10.
tRNA engineering for manipulating genetic code.
Katoh, T, Iwane, Y, Suga, H
RNA biology. 2018;(4-5):453-460
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Abstract
In ribosomal translation, only 20 kinds of proteinogenic amino acids (pAAs), namely 19 l-amino acids and glycine, are exclusively incorporated into polypeptide chain. To overcome this limitation, various methods to introduce non-proteinogenic amino acids (npAAs) other than the 20 pAAs have been developed to date. However, the repertoire of amino acids that can be simultaneously introduced is still limited. Moreover, the efficiency of npAA incorporation is not always sufficient depending on their structures. Fidelity of translation is sometimes low due to misincorporation of competing pAAs and/or undesired translation termination. Here, we provide an overview of efforts to solve these issues, focusing on the engineering of tRNAs.