Synthetic biology (Oxford, England)最新文献

{"title":"An injectable CRISPR therapy instructs B cells to produce anti-HIV antibodies.","authors":"Logan Thrasher Collins","doi":"10.1093/synbio/ysac027","DOIUrl":"https://doi.org/10.1093/synbio/ysac027","url":null,"abstract":"© The Author(s) 2022. Published by Oxford University Press. This is an Open Access article distributed under the terms of the Creative Commons Attribution-NonCommercial License (https://creativecommons.org/ licenses/by-nc/4.0/), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited. For commercial re-use, please contact journals.permissions@oup.com Although the immune system is well known as the guardian of the human body, certain infections and cancers can overwhelm its protective barriers. Over the past decade, scientists have developed genetic engineering tools that can enhance our immune systems to the point where they overcome such difficult threats. One success story in this area is the use of chimeric antigen receptor T cell (CAR T) therapy for blood cancers (1, 2). CAR T cells are engineered immune cells programmed to detect and destroy the cancer. In order to reprogram T cells, CAR T therapies require taking a blood sample out of a patient, shipping the sample to a laboratory, genetically modifying T cells within the sample, purifying the modified T cells, shipping them to the hospital and injecting them back into the patient. The cost, slowness and complexity of engineering immune cells outside of the body have limited accessibility of CAR T therapies and have challenged the expansion of this technology to the engineering of other immune cells such as B cells (3–5). To help overcome these barriers, a recent study was performed in Adi Barzel’s laboratory at Tel Aviv University and published in Nature Biotechnology. Nahmad et al. developed an injectable Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-based gene therapy to directly modify B cells inside of the body, giving them the ability to produce an antibody that fights acquired immune deficiency syndrome (AIDS) infections (6). In the future, such an injection might make immune cell therapies cheaper and thus more accessible to everyone and may pave the way for a vaccine against AIDS or a potent treatment for people who already suffer from the disease. CRISPR acts as a biomolecular cut-and-paste system that can insert genetic instructions at desired locations within the genome. It uses a protein–RNA complex consisting of a Cas9 protein and a guide RNA (gRNA) to cut a sequence within the genome that is recognized by the gRNA. After the cut has been made, one can provide a new piece of DNA instructions that the cell will stitch into the cut site during repair. CRISPR makes genetic alteration of cells much easier by precisely targeting where to put new DNA into the genome. Nahmad et al. injected mice with engineered adeno–associated viruses (AAVs) for delivery of (i) a gene encoding an anti–human immunodeficiency virus (HIV) antibody and (ii) genes encoding CRISPR Cas9 and gRNA machinery. AAVs act as a type of delivery system for transporting DNA into human cells and are commonly used in gene ther","PeriodicalId":74902,"journal":{"name":"Synthetic biology (Oxford, England)","volume":" ","pages":"ysac027"},"PeriodicalIF":0.0,"publicationDate":"2022-11-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9692189/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"40711918","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Piece by piece: making plant natural products accessible via heterologous biosynthesis.","authors":"Kira J Tiedge","doi":"10.1093/synbio/ysac028","DOIUrl":"https://doi.org/10.1093/synbio/ysac028","url":null,"abstract":"© The Author(s) 2022. Published by Oxford University Press. This is an Open Access article distributed under the terms of the Creative Commons Attribution-NonCommercial License (https://creativecommons.org/ licenses/by-nc/4.0/), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited. For commercial re-use, please contact journals.permissions@oup.com Numerous of the products we use in our daily lives originally come from plants. These plant-derived natural products include flavors, fragrances and medicines, such as the fragrance limonene from citrus fruits or the antimalarial drug artemisinin. As we cannot grow enough plants to satisfy our demand for these compounds, researchers are trying to manufacture these natural products in their laboratories via expression in easy-to-culture plants, bacteria or yeast—so-called heterologous hosts. Like this, researchers can create cell factories that can make more than what is made by the natural host. For smaller molecules like limonene, this is a fairly streamlined process as only one enzyme needs to be added to a host to create such a cell factory (1). However, for making chemically more complex bioactive molecules, adding 10 or more enzymatic reactions is required. Cloning these reactions being hard enough, the real bottleneck for making chemically complex natural products is the fact that the enzymes catalyzing the biosynthesis are often not known and need to be identified first. Recently, a group of researchers from the Max Planck Institute for Chemical Ecology in Jena, Germany, managed to decipher the multistep biosynthetic pathway of strychnine, a toxic alkaloid which is famously used as poison in crime stories and as a pesticide in real-world applications. Furthermore, they were able to transfer all required precursor enzymes as well as nine newly identified enzymes together into a heterologous host for transient expression (2), delivering a blueprint for creating cell factories able to perform complex plant chemistry. Hong et al.’s achievement is remarkable for several reasons: plants have very complex genomes, making it hard to identify which genes are encoding the biosynthesis of a desired product. For example, a plant genome can host many gene candidates that could act as the code for a specific enzymatic reaction. Finding out which is the right one involves a laborious screening process: the selected genes need to be cloned into an expression vector and then transformed into an expression system, such as tobacco plants (Nicotiana benthamiana), where their catalytic activity can be confirmed. Although advances in deoxyribonucleic acid (DNA) synthesis have helped in overcoming bottlenecks in cloning plant DNA (3), many challenges of identifying all the puzzle pieces that allow a plant to make a desired product and putting them together in the right order remain. The complex biosynthetic pathway of strychnine had puzzled the ","PeriodicalId":74902,"journal":{"name":"Synthetic biology (Oxford, England)","volume":" ","pages":"ysac028"},"PeriodicalIF":0.0,"publicationDate":"2022-11-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9700380/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"40504937","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"An arrayed CRISPR screen reveals Myc depletion to increase productivity of difficult-to-express complex antibodies in CHO cells.","authors":"Niels Bauer,&nbsp;Benedikt Oswald,&nbsp;Maximilian Eiche,&nbsp;Lisa Schiller,&nbsp;Emma Langguth,&nbsp;Christian Schantz,&nbsp;Andrea Osterlehner,&nbsp;Amy Shen,&nbsp;Shahram Misaghi,&nbsp;Julian Stingele,&nbsp;Simon Ausländer","doi":"10.1093/synbio/ysac026","DOIUrl":"https://doi.org/10.1093/synbio/ysac026","url":null,"abstract":"<p><p>Complex therapeutic antibody formats, such as bispecifics (bsAbs) or cytokine fusions, may provide new treatment options in diverse disease areas. However, the manufacturing yield of these complex antibody formats in Chinese Hamster Ovary (CHO) cells is lower than monoclonal antibodies due to challenges in expression levels and potential formation of side products. To overcome these limitations, we performed a clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR associated protein 9 (Cas9)-based knockout (KO) arrayed screening of 187 target genes in two CHO clones expressing two different complex antibody formats in a production-mimicking set-up. Our findings revealed that Myc depletion drastically increased product expression (>40%) by enhancing cell-specific productivity. The Myc-depleted cells displayed decreased cell densities together with substantially higher product titers in industrially-relevant bioprocesses using ambr15 and ambr250 bioreactors. Similar effects were observed across multiple different clones, each expressing a distinct complex antibody format. Our findings reinforce the mutually exclusive relationship between growth and production phenotypes and provide a targeted cell engineering approach to impact productivity without impairing product quality. We anticipate that CRISPR/Cas9-based CHO host cell engineering will transform our ability to increase manufacturing yield of high-value complex biotherapeutics.</p>","PeriodicalId":74902,"journal":{"name":"Synthetic biology (Oxford, England)","volume":" ","pages":"ysac026"},"PeriodicalIF":0.0,"publicationDate":"2022-11-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/02/0d/ysac026.PMC9700384.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"40504936","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"High-efficiency retron-mediated single-stranded DNA production in plants.","authors":"Wenjun Jiang,&nbsp;Gundra Sivakrishna Rao,&nbsp;Rashid Aman,&nbsp;Haroon Butt,&nbsp;Radwa Kamel,&nbsp;Khalid Sedeek,&nbsp;Magdy M Mahfouz","doi":"10.1093/synbio/ysac025","DOIUrl":"https://doi.org/10.1093/synbio/ysac025","url":null,"abstract":"<p><p>Retrons are a class of retroelements that produce multicopy single-stranded DNA (ssDNA) and participate in anti-phage defenses in bacteria. Retrons have been harnessed for the overproduction of ssDNA, genome engineering and directed evolution in bacteria, yeast and mammalian cells. Retron-mediated ssDNA production in plants could unlock their potential applications in plant biotechnology. For example, ssDNA can be used as a template for homology-directed repair (HDR) in several organisms. However, current gene editing technologies rely on the physical delivery of synthetic ssDNA, which limits their applications. Here, we demonstrated retron-mediated overproduction of ssDNA in <i>Nicotiana benthamiana</i>. Additionally, we tested different retron architectures for improved ssDNA production and identified a new retron architecture that resulted in greater ssDNA abundance. Furthermore, co-expression of the gene encoding the ssDNA-protecting protein VirE2 from <i>Agrobacterium tumefaciens</i> with the retron systems resulted in a 10.7-fold increase in ssDNA production <i>in vivo</i>. We also demonstrated clustered regularly interspaced short palindromic repeats-retron-coupled ssDNA overproduction and targeted HDR in <i>N. benthamiana</i>. Overall, we present an efficient approach for <i>in vivo</i> ssDNA production in plants, which can be harnessed for biotechnological applications. <b>Graphical Abstract</b>.</p>","PeriodicalId":74902,"journal":{"name":"Synthetic biology (Oxford, England)","volume":" ","pages":"ysac025"},"PeriodicalIF":0.0,"publicationDate":"2022-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/ab/6f/ysac025.PMC9700382.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"40504938","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Mouse chromosomes get supersized but find their limits.","authors":"David M Truong","doi":"10.1093/synbio/ysac024","DOIUrl":"https://doi.org/10.1093/synbio/ysac024","url":null,"abstract":"© The Author(s) 2022. Published by Oxford University Press. This is an Open Access article distributed under the terms of the Creative Commons Attribution-NonCommercial License (https://creativecommons.org/ licenses/by-nc/4.0/), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited. For commercial re-use, please contact journals.permissions@oup.com Humans diverged from primates when an ancestral chromosomal fusion—the event when two chromosomes join together to form one—gave us 23 instead of 24 sets of chromosomes. In muntjac deer, small deer native to South and Southeast Asia, chromosome fusions occurred so often that Indian muntjacs have only 3 chromosomes, whereas Chinese muntjacs have 23 (1). Fusions matter not only during the evolution of species but can also cause diseases such as cancer or Down’s syndrome. While fusions occur often in nature, engineering events like these on purpose have been difficult to do. The field of synthetic genomics attempts feats like this, along with building new designer chromosomes for applications in medicine, agriculture and industrial processing. Completely synthetic genomes have been built for bacteria (2), as well as for yeast (2). Additionally, it was shown that all 16 yeast chromosomes can be fused into one single chromosome 12 megabases long (3). Besides these achievements, it remained an open question what the actual size limit of a single chromosome would be, for example, whether the 100–200 megabase mammalian chromosomes could be fused and whether changes like these would persist through multiple generations. Answers could be used to model speciation and human diseases, as well as biologically ‘contain’ engineered organisms from natural populations. In a groundbreaking new study (4), researchers from the Chinese Academy of Sciences have generated the largest designed fusion chromosomes so far reported in mice as their research model. To technically achieve this, they used Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) to make targeted DNA breaks. These breaks would induce recombination—a natural repair process of the cell—thereby fusing the two largest mouse chromosomes 1 and 2 into a single one, in two different orientations, followed by fusing medium size chromosomes 4 and 5. The longest of these chromosomes was 377 megabases long and functional. In addition, they accomplished all this engineering in haploid mouse embryonic stem cells (i.e. cells with only one set of chromosomes), showing the potential to make mice easier to engineer using haploid cells in a Petri dish. Although the authors could generate heterozygous embryos with the largest fused chromosomes, one fused orientation was lethal to the developing embryo, while embryos of the other orientation grew to adulthood. Yet, the resultant mice could not breed homozygous offspring. Surprisingly, the 308 megabase medium-sized fused chromosome mice coul","PeriodicalId":74902,"journal":{"name":"Synthetic biology (Oxford, England)","volume":" ","pages":"ysac024"},"PeriodicalIF":0.0,"publicationDate":"2022-10-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/19/9b/ysac024.PMC9659764.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"40687767","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"basicsynbio and the BASIC SEVA collection: software and vectors for an established DNA assembly method.","authors":"Matthew C Haines,&nbsp;Benedict Carling,&nbsp;James Marshall,&nbsp;Vasily A Shenshin,&nbsp;Geoff S Baldwin,&nbsp;Paul Freemont,&nbsp;Marko Storch","doi":"10.1093/synbio/ysac023","DOIUrl":"https://doi.org/10.1093/synbio/ysac023","url":null,"abstract":"<p><p>Standardized deoxyribonucleic acid (DNA) assembly methods utilizing modular components provide a powerful framework to explore designs and iterate through Design-Build-Test-Learn cycles. Biopart Assembly Standard for Idempotent Cloning (BASIC) DNA assembly uses modular parts and linkers, is highly accurate, easy to automate, free for academic and commercial use and enables hierarchical assemblies through an idempotent format. These features enable applications including pathway engineering, ribosome binding site (RBS) tuning, fusion protein engineering and multiplexed guide ribonucleic acid (RNA) expression. In this work, we present basicsynbio, open-source software encompassing a Web App (https://basicsynbio.web.app/) and Python Package (https://github.com/LondonBiofoundry/basicsynbio), enabling BASIC construct design via simple drag-and-drop operations or programmatically. With basicsynbio, users can access commonly used BASIC parts and linkers while designing new parts and assemblies with exception handling for common errors. Users can export sequence data and create instructions for manual or acoustic liquid-handling platforms. Instruction generation relies on the BasicBuild Open Standard, which is parsed for bespoke workflows and is serializable in JavaScript Object Notation for transfer and storage. We demonstrate basicsynbio, assembling 30 vectors using sequences including modules from the Standard European Vector Architecture (SEVA). The BASIC SEVA vector collection is compatible with BASIC and Golden Gate using BsaI. Vectors contain one of six antibiotic resistance markers and five origins of replication from different compatibility groups. The collection is available via Addgene under an OpenMTA agreement. Furthermore, vector sequences are available from within the basicsynbio application programming interface with other collections of parts and linkers, providing a powerful environment for designing assemblies for bioengineering applications. <b>Graphical Abstract</b>.</p>","PeriodicalId":74902,"journal":{"name":"Synthetic biology (Oxford, England)","volume":" ","pages":"ysac023"},"PeriodicalIF":0.0,"publicationDate":"2022-10-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9664905/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"40687691","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Could a simple model of COVID-19 infections be the key to designing better virus-based therapies?","authors":"Connor R King","doi":"10.1093/synbio/ysac019","DOIUrl":"https://doi.org/10.1093/synbio/ysac019","url":null,"abstract":"© The Author(s) 2022. Published by Oxford University Press. This is an Open Access article distributed under the terms of the Creative Commons Attribution-NonCommercial License (https://creativecommons.org/licenses/by-nc/4.0/), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited. For commercial re-use, please contact journals.permissions@oup.com Since the emergence of COVID-19, the spotlight on viruses has become negative. However, viruses are not only our enemies, due to their ability to deliver DNA and RNA into cells, viruses can also be repurposed as therapeutics. This ability is already used to treat genetic diseases and cancer (2, 4). However, an important factor that is often overlooked when developing virus-based therapies is the fact that each patient’s immune system might respond differently to the treatment which can determine its effectiveness (3). Recently, the Ke Lab at Los Alamos National Laboratory developed a simplistic model of the immune response to SARS-CoV-2 infection that can explain why some individuals have severe symptoms while others quickly resolve the infection (1). The simplicity of this model suggests that one day it could not only help understand viral infections but help improve virus-based therapies. Current designs for virus-based therapeutics mainly focused on the delivery system itself. Much like if you were designing a system to deliver medicine to houses, you might first want to optimize the delivery vehicle to be used for delivering the medicine and identify routes for the delivery. Now imagine that this system was developed just around the delivery of the medication itself without consideration of what might happen when the medicine is delivered. If you deliver this medicine to a house that is, much like a cell, unaware of why the medicine is coming, the recipient may dispose of the said medication. Since the injection of DNA or RNA from a virus into a cell is typically associated with a disease, it is reasonable to assume that the cell has processes at hand that interfere with virus-based treatments. The model from the Ke lab might help therapy-designers to predict and mitigate these processes just as the model is able to explain why some people get severe COVID-19 and others do not. The focus of the model is the immune response generated by the molecular footprint left by the viral disease. This footprint results from the viral infection itself but is also generated by cells being damaged by the immune system. Both contribute to sustaining an active immune response in patients. The model simplifies many of the processes that generate and remove this footprint in order to reduce complexity. Their approach to making the model simple can be compared to trying to plan out how long it takes to drive from San Francisco to Los Angeles. There are many factors that contribute to how quickly one can drive—traffic and headwind—but one could p","PeriodicalId":74902,"journal":{"name":"Synthetic biology (Oxford, England)","volume":" ","pages":"ysac019"},"PeriodicalIF":0.0,"publicationDate":"2022-09-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/07/5c/ysac019.PMC9518665.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"40390049","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A toolkit for enhanced reproducibility of RNASeq analysis for synthetic biologists.","authors":"Benjamin J Garcia,&nbsp;Joshua Urrutia,&nbsp;George Zheng,&nbsp;Diveena Becker,&nbsp;Carolyn Corbet,&nbsp;Paul Maschhoff,&nbsp;Alexander Cristofaro,&nbsp;Niall Gaffney,&nbsp;Matthew Vaughn,&nbsp;Uma Saxena,&nbsp;Yi-Pei Chen,&nbsp;D Benjamin Gordon,&nbsp;Mohammed Eslami","doi":"10.1093/synbio/ysac012","DOIUrl":"https://doi.org/10.1093/synbio/ysac012","url":null,"abstract":"<p><p>Sequencing technologies, in particular RNASeq, have become critical tools in the design, build, test and learn cycle of synthetic biology. They provide a better understanding of synthetic designs, and they help identify ways to improve and select designs. While these data are beneficial to design, their collection and analysis is a complex, multistep process that has implications on both discovery and reproducibility of experiments. Additionally, tool parameters, experimental metadata, normalization of data and standardization of file formats present challenges that are computationally intensive. This calls for high-throughput pipelines expressly designed to handle the combinatorial and longitudinal nature of synthetic biology. In this paper, we present a pipeline to maximize the analytical reproducibility of RNASeq for synthetic biologists. We also explore the impact of reproducibility on the validation of machine learning models. We present the design of a pipeline that combines traditional RNASeq data processing tools with structured metadata tracking to allow for the exploration of the combinatorial design in a high-throughput and reproducible manner. We then demonstrate utility via two different experiments: a control comparison experiment and a machine learning model experiment. The first experiment compares datasets collected from identical biological controls across multiple days for two different organisms. It shows that a reproducible experimental protocol for one organism does not guarantee reproducibility in another. The second experiment quantifies the differences in experimental runs from multiple perspectives. It shows that the lack of reproducibility from these different perspectives can place an upper bound on the validation of machine learning models trained on RNASeq data. Graphical Abstract.</p>","PeriodicalId":74902,"journal":{"name":"Synthetic biology (Oxford, England)","volume":" ","pages":"ysac012"},"PeriodicalIF":0.0,"publicationDate":"2022-08-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/4f/a8/ysac012.PMC9408027.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"33444691","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A universal approach to gene expression engineering.","authors":"Rahmi Lale,&nbsp;Lisa Tietze,&nbsp;Maxime Fages-Lartaud,&nbsp;Jenny Nesje,&nbsp;Ingerid Onsager,&nbsp;Kerstin Engelhardt,&nbsp;Che Fai Alex Wong,&nbsp;Madina Akan,&nbsp;Niklas Hummel,&nbsp;Jörn Kalinowski,&nbsp;Christian Rückert,&nbsp;Martin Frank Hohmann-Marriott","doi":"10.1093/synbio/ysac017","DOIUrl":"https://doi.org/10.1093/synbio/ysac017","url":null,"abstract":"<p><p>In this study, we provide a universal approach to Gene Expression Engineering (GeneEE) for creating artificial expression systems. GeneEE leads to the generation of artificial 5^' regulatory sequences (ARES) consisting of promoters and 5^' untranslated regions. The ARES lead to the successful recruitment of RNA polymerase, related sigma factors and ribosomal proteins that result in a wide range of expression levels. We also demonstrate that by engaging native transcription regulators, GeneEE can be used to generate inducible promoters. To showcase the universality of the approach, we demonstrate that 200-nucleotide (nt)-long DNA with random composition can be used to generate functional expression systems in six bacterial species, <i>Escherichia coli, Pseudomonas putida, Corynebacterium glutamicum, Thermus thermophilus, Streptomyces albus</i> and <i>Streptomyces lividans</i>, and the eukaryote yeast <i>Saccharomyces cerevisiae</i>.</p>","PeriodicalId":74902,"journal":{"name":"Synthetic biology (Oxford, England)","volume":" ","pages":"ysac017"},"PeriodicalIF":0.0,"publicationDate":"2022-08-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/f4/c8/ysac017.PMC9534286.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"33497337","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Evaluating the persistence and stability of a DNA-barcoded microbial system in a mock home environment.","authors":"Nathan D McDonald,&nbsp;Katherine A Rhea,&nbsp;John P Davies,&nbsp;Julie L Zacharko,&nbsp;Kimberly L Berk,&nbsp;Patricia E Buckley","doi":"10.1093/synbio/ysac016","DOIUrl":"https://doi.org/10.1093/synbio/ysac016","url":null,"abstract":"<p><p>Recent advancements in engineered microbial systems capable of deployment in complex environments have enabled the creation of unique signatures for environmental forensics operations. These microbial systems must be robust, able to thrive in specific environments of interest and contain molecular signatures, enabling the detection of the community across conditions. Furthermore, these systems must balance biocontainment concerns with the stability and persistence required for environmental forensics. Here we evaluate the stability and persistence of a recently described microbial system composed of germination-deficient <i>Bacillus subtilis</i> and <i>Saccharomyces cerevisiae</i> spores containing nonredundant DNA barcodes in a controlled simulated home environment. These spore-based microbial communities were found to be persistent in the simulated environment across 30-day periods and across multiple surface types. To improve the repeatability and reproducibility in detecting the DNA barcodes, we evaluated several spore lysis and sampling processes paired with Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) -CRISPR-associated proteins (Cas) detection (Sherlock). Finally, having optimized the detectability of the spores, we demonstrate that we can detect the spores transferring across multiple material types. Together, we further demonstrate the utility of a recently described microbial forensics system and highlight the importance of independent validation and verification of synthetic biology tools and applications. Graphical Abstract.</p>","PeriodicalId":74902,"journal":{"name":"Synthetic biology (Oxford, England)","volume":" ","pages":"ysac016"},"PeriodicalIF":0.0,"publicationDate":"2022-08-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9423098/pdf/ysac016.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"40335395","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}