Synthetic biology (Oxford, England)最新文献

{"title":"Periplasmic stress contributes to a trade-off between protein secretion and cell growth in <i>Escherichia coli</i> Nissle 1917.","authors":"Sivaram Subaya Emani,&nbsp;Anton Kan,&nbsp;Timothy Storms,&nbsp;Shanna Bonanno,&nbsp;Jade Law,&nbsp;Sanhita Ray,&nbsp;Neel S Joshi","doi":"10.1093/synbio/ysad013","DOIUrl":"https://doi.org/10.1093/synbio/ysad013","url":null,"abstract":"<p><p>Maximizing protein secretion is an important target in the design of engineered living systems. In this paper, we characterize a trade-off between cell growth and per-cell protein secretion in the curli biofilm secretion system of <i>Escherichia coli</i> Nissle 1917. Initial characterization using 24-h continuous growth and protein production monitoring confirms decreased growth rates at high induction, leading to a local maximum in total protein production at intermediate induction. Propidium iodide (PI) staining at the endpoint indicates that cellular death is a dominant cause of growth reduction. Assaying variants with combinatorial constructs of inner and outer membrane secretion tags, we find that diminished growth at high production is specific to secretory variants associated with periplasmic stress mediated by outer membrane secretion and periplasmic accumulation of protein containing the outer membrane transport tag. RNA sequencing experiments indicate upregulation of known periplasmic stress response genes in the highly secreting variant, further implicating periplasmic stress in the growth-secretion trade-off. Overall, these results motivate additional strategies for optimizing total protein production and longevity of secretory engineered living systems <b>Graphical Abstract</b>.</p>","PeriodicalId":74902,"journal":{"name":"Synthetic biology (Oxford, England)","volume":"8 1","pages":"ysad013"},"PeriodicalIF":0.0,"publicationDate":"2023-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/c2/ae/ysad013.PMC10439730.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10199999","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":"Robustness and reproducibility of simple and complex synthetic logic circuit designs using a DBTL loop.","authors":"Breschine Cummins,&nbsp;Justin Vrana,&nbsp;Robert C Moseley,&nbsp;Hamed Eramian,&nbsp;Anastasia Deckard,&nbsp;Pedro Fontanarrosa,&nbsp;Daniel Bryce,&nbsp;Mark Weston,&nbsp;George Zheng,&nbsp;Joshua Nowak,&nbsp;Francis C Motta,&nbsp;Mohammed Eslami,&nbsp;Kara Layne Johnson,&nbsp;Robert P Goldman,&nbsp;Chris J Myers,&nbsp;Tessa Johnson,&nbsp;Matthew W Vaughn,&nbsp;Niall Gaffney,&nbsp;Joshua Urrutia,&nbsp;Shweta Gopaulakrishnan,&nbsp;Vanessa Biggers,&nbsp;Trissha R Higa,&nbsp;Lorraine A Mosqueda,&nbsp;Marcio Gameiro,&nbsp;Tomáš Gedeon,&nbsp;Konstantin Mischaikow,&nbsp;Jacob Beal,&nbsp;Bryan Bartley,&nbsp;Tom Mitchell,&nbsp;Tramy T Nguyen,&nbsp;Nicholas Roehner,&nbsp;Steven B Haase","doi":"10.1093/synbio/ysad005","DOIUrl":"https://doi.org/10.1093/synbio/ysad005","url":null,"abstract":"<p><p>Computational tools addressing various components of design-build-test-learn (DBTL) loops for the construction of synthetic genetic networks exist but do not generally cover the entire DBTL loop. This manuscript introduces an end-to-end sequence of tools that together form a DBTL loop called Design Assemble Round Trip (DART). DART provides rational selection and refinement of genetic parts to construct and test a circuit. Computational support for experimental process, metadata management, standardized data collection and reproducible data analysis is provided via the previously published Round Trip (RT) test-learn loop. The primary focus of this work is on the Design Assemble (DA) part of the tool chain, which improves on previous techniques by screening up to thousands of network topologies for robust performance using a novel robustness score derived from dynamical behavior based on circuit topology only. In addition, novel experimental support software is introduced for the assembly of genetic circuits. A complete design-through-analysis sequence is presented using several OR and NOR circuit designs, with and without structural redundancy, that are implemented in budding yeast. The execution of DART tested the predictions of the design tools, specifically with regard to robust and reproducible performance under different experimental conditions. The data analysis depended on a novel application of machine learning techniques to segment bimodal flow cytometry distributions. Evidence is presented that, in some cases, a more complex build may impart more robustness and reproducibility across experimental conditions. <b>Graphical Abstract</b>.</p>","PeriodicalId":74902,"journal":{"name":"Synthetic biology (Oxford, England)","volume":"8 1","pages":"ysad005"},"PeriodicalIF":0.0,"publicationDate":"2023-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/5b/51/ysad005.PMC10105856.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9736862","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":"Engineered microbes report levels of freshwater contamination in real time.","authors":"Tea Crnković","doi":"10.1093/synbio/ysad002","DOIUrl":"https://doi.org/10.1093/synbio/ysad002","url":null,"abstract":"","PeriodicalId":74902,"journal":{"name":"Synthetic biology (Oxford, England)","volume":"8 1","pages":"ysad002"},"PeriodicalIF":0.0,"publicationDate":"2023-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9969829/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10820814","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 economy of details: standards and data reusability.","authors":"Ana Delgado","doi":"10.1093/synbio/ysac030","DOIUrl":"https://doi.org/10.1093/synbio/ysac030","url":null,"abstract":"<p><p>Reusability has been a key issue since the origins of the parts-based approach to synthetic biology. Starting with the BioBrick™ standard part, multiple efforts have aimed to make biology more exchangeable. The reusability of parts and other deoxyribonucleic acid-based data has proven over time to be challenging, however. Drawing on a series of qualitative interviews and an international workshop, this article explores the challenges of reusability in real laboratory practice. It shows particular ways that standards are experienced as presenting shortcomings for capturing the kinds of contextual information crucial for scientists to be able to reuse biological parts and data. I argue that researchers in specific laboratories develop a sense of how much circumstantial detail they need to share for others to be able to make sense of their data and possibly reuse it. When choosing particular reporting formats, recharacterizing data to gain closer knowledge or requesting additional information, researchers enact an 'economy of details'. The farther apart two laboratories are in disciplinary, epistemological, technical and geographical terms, the more detailed information needs to be captured for data to be reusable across contexts. In synthetic biology, disciplinary distance between computing science and engineering researchers and experimentalist biologists is reflected in diverging views on standards: what kind of information should be included to enable reusability, what kind of information can be captured by standards at all and how they may serve to produce and circulate knowledge. I argue that such interdisciplinary tensions lie at the core of difficulties in setting standards in synthetic biology.</p>","PeriodicalId":74902,"journal":{"name":"Synthetic biology (Oxford, England)","volume":"8 1","pages":"ysac030"},"PeriodicalIF":0.0,"publicationDate":"2023-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/e7/47/ysac030.PMC9817096.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10518569","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":"Variability in genome-engineering source materials: consider your starting point.","authors":"Simona Patange,&nbsp;Sierra D Miller,&nbsp;Samantha D Maragh","doi":"10.1093/synbio/ysad003","DOIUrl":"https://doi.org/10.1093/synbio/ysad003","url":null,"abstract":"<p><p>The presence and impact of variability in cells as the source material for genome engineering are important to consider for the design, execution and interpretation of outcomes of a genome-engineering process. Variability may be present at the genotype and phenotype level, yet the impact of these sources of variability on a genome-engineering experiment may not be regularly considered by researchers. In this perspective, we use clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated protein (Cas) genome editing of mammalian cells to provide examples of how variation within or across cell samples may mislead a researcher in their expectations about the cells they are engineering. Furthermore, we highlight the need for understanding the baseline cell genotype and phenotype to appropriately understand the starting cell material and interpret and attribute the impact of engineering on cells. We emphasize that heterogeneity within a cell pool and the inherent variability in the cellular materials used for genome engineering are complex, but of high value to characterize and account for where possible, to move toward the potential of generating desired and predictable engineered products. Provided is a framework cause-and-effect diagram for CRISPR/Cas9 genome editing toward identifying and mitigating potential sources of variability. We encourage researchers to consider the variability of source materials and undertake strategies, which may include those described here, for detecting, attributing and minimizing additional sources of variability where possible toward the aim of fostering greater reliability, confidence and reproducibility in genome-engineering studies. <b>Graphical Abstract</b>.</p>","PeriodicalId":74902,"journal":{"name":"Synthetic biology (Oxford, England)","volume":"8 1","pages":"ysad003"},"PeriodicalIF":0.0,"publicationDate":"2023-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10029982/pdf/ysad003.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9172013","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":"Fighting fire with fire: engineering a microbe into a therapeutic defense against drug-resistant biofilms.","authors":"Charlotte Ayn Cialek","doi":"10.1093/synbio/ysad008","DOIUrl":"https://doi.org/10.1093/synbio/ysad008","url":null,"abstract":"","PeriodicalId":74902,"journal":{"name":"Synthetic biology (Oxford, England)","volume":"8 1","pages":"ysad008"},"PeriodicalIF":0.0,"publicationDate":"2023-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10171107/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9839062","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 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}