Journal of Biological Engineering最新文献

{"title":"Genetically engineered secretory horseradish peroxidase is a sensitive, stable, and affordable non-lytic reporter gene system for real-time promoter activity management.","authors":"Mu-Shen Chang, Chia-Yi Lee, Yu-Yen Chang, Pin-Jie Li, Hsin-Yu Wu, En-Shuo Liu, Hsin-Kai Huang, Wen-Wei Lin","doi":"10.1186/s13036-025-00508-w","DOIUrl":"https://doi.org/10.1186/s13036-025-00508-w","url":null,"abstract":"<p><p>A light-producing secretory protein that is collectible through the supernatant of a culture medium is essential in a cell-based reporter gene system and allows for real-time monitoring of upstream events of a promoter. Compared to other secretory luciferases, Cypridina luciferase (CLuc) coupled with vargulin emits the brightest signal; however, the signal half-life suffers constantly from the fast oxidation process of the substrate, resulting in a rapid signal depletion, which makes the detection signal short and unstable. In this study, we aimed to develop a new reporter gene system with a more stable signal and lower cost, whilst retaining sensitivity comparable to the CLuc reporter gene system. To this end, we genetically engineered horseradish peroxidase (HRP) to be secreted with mammalian cells. The secreted form HRP (sHRP) was then used as a proof-of-concept of real-time cell signaling management. First, we made sure that HRP retained its enzymatic function with our genetic engineering process and confirmed that it was collectable and suitable for side-by-side comparison with CLuc. sHRP showed comparable sensitivity, 7 to 80 times more signal half-life compared to CLuc, and precisely reported NF-κB-regulated promoter in response to stimulation with TNF-α. sHRP was not affected by multiple cell culturing media and was calculated to be at least 9 times cheaper than the CLuc reporter gene system. Thus, sHRP offers new insight into the reporter gene system for drug screening and intracellular signaling management and provides a precise, sustainable and affordable operating environment.</p>","PeriodicalId":15053,"journal":{"name":"Journal of Biological Engineering","volume":"19 1","pages":"37"},"PeriodicalIF":5.7,"publicationDate":"2025-04-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12012966/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143998476","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Decellularized apple hypanthium as a plant-based biomaterial for cartilage regeneration in vitro: a comparative study of progenitor cell types and environmental conditions.","authors":"Mira Hammad, Justin Dugué, Eric Maubert, Catherine Baugé, Karim Boumédiene","doi":"10.1186/s13036-025-00502-2","DOIUrl":"https://doi.org/10.1186/s13036-025-00502-2","url":null,"abstract":"<p><strong>Background: </strong>Decellularized plant tissues have been shown to enhance the integration and proliferation of human cells, demonstrating biocompatibility. These tissues are now being considered as valuable biomaterials for tissue engineering due to their diverse architectures and favorable cytocompatibility. In this study, we assessed decellularized apple hypanthium as a potential biomaterial for generating cartilage-like structures, utilizing four different progenitor cell types and varying environmental conditions in vitro.</p><p><strong>Results: </strong>Cell viability assays indicated integration and cell proliferation. Histological staining and gene expression analyses confirmed the synthesis and deposition of a cartilaginous extracellular matrix. Notably, hypoxia had varying effects on chondrogenesis based on the cell type. Among the progenitor cells evaluated, those derived from auricular perichondrium were particularly promising, as they differentiated into chondrocytes without requiring a low-oxygen environment. Additionally, our findings demonstrated that apple-derived biomaterials outperformed microencapsulation in alginate beads in promoting chondrogenesis.</p><p><strong>Conclusion: </strong>These results highlight the potential of plant-based biomaterials for the development of implantable devices for cartilage regeneration and suggest broader applications in tissue engineering and future clinical endeavors.</p>","PeriodicalId":15053,"journal":{"name":"Journal of Biological Engineering","volume":"19 1","pages":"38"},"PeriodicalIF":5.7,"publicationDate":"2025-04-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12012941/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144010755","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Anti-inflammatory activity of magnetic fields emitted by graphene devices on cultured human cells.","authors":"Sara Franceschelli, Federica De Cecco, Stefano Benedetti, Valeria Panella, Lorenza Speranza, Alfredo Grilli, Pierdomenico D'Andrea","doi":"10.1186/s13036-025-00507-x","DOIUrl":"https://doi.org/10.1186/s13036-025-00507-x","url":null,"abstract":"<p><strong>Background: </strong>Inflammation plays a key role in various diseases such as pancreatitis, cancer, and rheumatoid arthritis. Acute inflammation involves processes like vasodilation, increased vascular permeability, and leukocyte accumulation, which lead to cellular damage due to reactive oxygen species (ROS). Low-frequency electromagnetic fields (ELF-EMFs) have shown potential in reducing oxidative stress and inflammation. This study assesses the effectiveness of a new wearable device containing graphene quantum dots in reducing inflammation and oxidative stress in Jurkat T cells stimulated by lipopolysaccharide (LPS). The device is evaluated for its impact on ROS production and inflammation.</p><p><strong>Results: </strong>The results show that the device significantly lowers ROS levels and reduces the inflammatory response by decreasing pro-inflammatory cytokines such as IL-6, TNF-α, and IL-1β. Additionally, the device inhibits LPS-induced iNOS and COX-2 activity and modulates NF-κB signaling, indicating its potential as a therapeutic tool for managing inflammation and oxidative stress.</p><p><strong>Conclusion: </strong>These findings highlight the device's ability to combat inflammation, offering a non-invasive and effective approach for inflammatory diseases.</p>","PeriodicalId":15053,"journal":{"name":"Journal of Biological Engineering","volume":"19 1","pages":"36"},"PeriodicalIF":5.7,"publicationDate":"2025-04-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12013070/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144006699","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Engineering FcRn binding kinetics dramatically extends antibody serum half-life and enhances therapeutic potential.","authors":"Sanghwan Ko, Migyeong Jo, Munsu Kyung, Wonju Lee, Woo Hyung Ko, Jung-Hyun Na, Youn Seo Chun, Byoung Joon Ko, Sang Taek Jung","doi":"10.1186/s13036-025-00506-y","DOIUrl":"https://doi.org/10.1186/s13036-025-00506-y","url":null,"abstract":"<p><strong>Background: </strong>Optimizing the IgG Fc domain for neonatal Fc receptor (FcRn) binding is crucial for enhancing antibody pharmacokinetics. The prolonged serum half-life of IgG antibody is governed by its pH-dependent interaction with FcRn, enabling efficient binding at acidic endosomal pH, intracellular trafficking, and release at neutral serum pH. However, a critical yet previously unrecognized challenge in Fc engineering for extending the serum half-life of therapeutic antibodies is the intense competition with endogenous IgG for FcRn binding during intracellular trafficking, which limits FcRn-mediated transport and reduces the serum persistence of therapeutic antibodies. To address this, we developed an Fc variant that precisely modulates pH-dependent FcRn binding kinetics, accelerates FcRn association at acidic pH, and promotes rapid dissociation at neutral pH, thereby enhancing FcRn-driven intracellular transport, outcompeting endogenous IgG, and achieving unprecedented improvement in the serum half-life of therapeutic antibodies.</p><p><strong>Results: </strong>Using comprehensive site-directed saturation mutagenesis coupled with functional screening, we generated a diverse panel of Fc variants and identified two with distinct FcRn binding kinetics: YML (L309Y/Q311M/M428L), which exhibited superior FcRn association at acidic pH and accelerated dissociation at neutral pH, and EML (L309E/Q311M/M428L), which displayed attenuated binding kinetics. In human FcRn transgenic mice, YML extended the serum half-life of clinically used trastuzumab with a wild-type Fc by 6.1-fold, demonstrating a remarkable improvement over previously reported Fc-engineered variants, including PFc29 (Q311R/M428L) and DHS (L309D/Q311H/N434S), which represent the most effective Fc modifications for prolonging serum persistence to date. This in vivo validation underscores the pivotal role of FcRn kinetic tuning in overcoming endogenous IgG competition and maximizing FcRn-mediated antibody transport. Additionally, YML exhibited potent complement-dependent cytotoxicity (CDC) while maintaining favorable physicochemical properties.</p><p><strong>Conclusion: </strong>This study presents a rational Fc engineering framework to optimize FcRn binding kinetics, addressing a previously unconsidered challenge-endogenous IgG competition during intracellular trafficking of therapeutic antibodies. The distinct kinetic behaviors of YML and EML highlight the critical necessity of precise control over pH-dependent association and dissociation rates in FcRn binding. YML represents a next-generation Fc platform, offering enhanced pharmacokinetics and improved effector functions, thus providing a powerful strategy for developing biologics with superior serum persistence and therapeutic efficacy.</p>","PeriodicalId":15053,"journal":{"name":"Journal of Biological Engineering","volume":"19 1","pages":"35"},"PeriodicalIF":5.7,"publicationDate":"2025-04-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12007268/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144000734","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Correction: Microfluidic device for enhancement and analysis of osteoblast differentiation in three-dimensional cell cultures.","authors":"Michael Killinger, Adéla Kratochvilová, Eva Ingeborg Reihs, Eva Matalová, Karel Klepárník, Mario Rothbauer","doi":"10.1186/s13036-025-00501-3","DOIUrl":"https://doi.org/10.1186/s13036-025-00501-3","url":null,"abstract":"","PeriodicalId":15053,"journal":{"name":"Journal of Biological Engineering","volume":"19 1","pages":"33"},"PeriodicalIF":5.7,"publicationDate":"2025-04-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12004551/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143966804","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Unraveling the unique bioactivities of highly purified C-phycocyanin and allophycocyanin.","authors":"Jimin Na, Soobin Jang, Myeongkwan Song, SeungEun Nam, Woon-Yong Choi, Hwasung Shin, Soonjo Kwon, Youngbin Baek","doi":"10.1186/s13036-025-00496-x","DOIUrl":"https://doi.org/10.1186/s13036-025-00496-x","url":null,"abstract":"<p><strong>Background: </strong>The blue-green microalgae Spirulina, used in human nutrition for centuries, includes phycobiliproteins such as C-phycocyanin (CPC) and allophycocyanin (APC). Assessing their unique bioactivities separately is difficult as they have similar properties, such as molecular weight and isoelectric point. In the present study, we aimed to separate CPC and APC and to evaluate their bioactivities. CPC and APC were separated using a hydrophobic membrane and ammonium sulfate, which promotes reversible and specific protein binding to the membrane. Spectroscopic analysis, HPLC, and SDS-PAGE revealed a successful separation of CPC and APC. Their bioactivities were evaluated through CCK- 8 assays for anticancer activity, radical scavenging assays for antioxidant activity, and albumin denaturation assays for anti-inflammatory activity, respectively.</p><p><strong>Results: </strong>The results revealed that highly purified APC showed 40% higher anticancer activity than the control, whereas CPC increased the viability of cancer cells, resulting in a 30% decrease in anticancer activity compared to the control. In contrast, highly purified CPC showed approximately 25% higher antioxidant activity and twice as much anti-inflammatory activity as APCs; moreover, the presence of both showed higher antioxidant activity.</p><p><strong>Conclusion: </strong>This study provides important insights into the unique bioactivities of CPC and APC for their appropriate application as anticancer, antiphlogistic, and antioxidant agents.</p>","PeriodicalId":15053,"journal":{"name":"Journal of Biological Engineering","volume":"19 1","pages":"34"},"PeriodicalIF":5.7,"publicationDate":"2025-04-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12004856/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144010760","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Unsilencing a cryptic xylose metabolic pathway in Rhodococcus jostii RHA1 for efficient lipid production from lignocellulosic biomass.","authors":"Miguel G Acedos, Isabel De la Torre, Jorge Barriuso, José L García","doi":"10.1186/s13036-025-00503-1","DOIUrl":"https://doi.org/10.1186/s13036-025-00503-1","url":null,"abstract":"<p><p>Rhodococcus jostii RHA1 is an oleaginous bacterium that has attracted considerable attention due to its capacity to use different carbon sources to accumulate significant levels of triacylglycerols that might be converted into biofuels. However, this strain cannot transform xylose into lipids reducing its potential when growing on saccharified lignocellulosic biomass. In this work, we demonstrate that wild type R. jostii RHA1 can be evolved by adaptive laboratory evolution (ALE) to metabolize xylose without engineering heterologous metabolic pathways in the host. We have generated a phenotypically adapted ALE-xyl strain able to use xylose as the sole carbon and energy source more efficiently that an engineered recombinant strain harbouring heterologous xylA and xylB genes encoding a xylose isomerase metabolic pathway. The R. jostii RHA1 ALE-xyl strain accumulates lipids very efficiently using xylose as substrate, but even more importantly it can consume glucose and xylose at the same time. Transcriptomic analyses of ALE-xyl strain growing with glucose or xylose revealed the existence of a silent pentose metabolizing operon that is overexpressed in the presence of xylose. The detection of a xylose reductase activity together with the presence of xylitol in the cytoplasm of ALE-xyl strain suggests that xylose is consumed by a reductase pathway. This study demonstrates that, in cases where a clear phenotypic selection method is available, ALE can be used to improve very efficiently industrial microbial strains without using genetic engineering tools. Strategies focused to exploit the silent phenotypic flexibility of microorganisms to metabolize different carbon sources are powerful tools for the production of microbial value-added products using saccharified lignocellulosic wastes.</p>","PeriodicalId":15053,"journal":{"name":"Journal of Biological Engineering","volume":"19 1","pages":"32"},"PeriodicalIF":5.7,"publicationDate":"2025-04-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11998424/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144025525","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Biomaterial-based chitosan nanohydrogel films: combination of Bistorta officinalis and Ca-doped carbon dots for improved blood clotting.","authors":"Hassan Tavakoli, Meysam Najaflou, Ahmad Yarikhosroushahi","doi":"10.1186/s13036-025-00498-9","DOIUrl":"https://doi.org/10.1186/s13036-025-00498-9","url":null,"abstract":"<p><strong>Background: </strong>Bleeding and traumatic injuries are still a major issue necessitating the development of advanced hemostatic materials that are economical, biocompatible, and effective. Chitosan's (CS) haemostatic and biocompatible properties make it a promising wound-healing material, however, effective cross-linking is essential for appropriate physiochemical properties. In this study, calcium-doped carbon dots (CDs) produced from coriander leaves were used as cross-linking agents to improve the functional performance and structural integrity of nanohydrogel films. Furthermore, extract of the medicinal plant Bistorta officinalis (BEX), a traditional medicinal plant with strong hemostatic and antibacterial qualities, was incorporated into the hydrogel matrix.</p><p><strong>Results: </strong>Analysis and characterization of the synthesized CDs thoroughly confirmed that they have monodispersed spherical shape, negative zeta potential, and active functional groups which effectively cross-linked the chitosan matrix and increased the mechanical strength and stability of the film. Cytotoxicity and antibacterial results of the final films showed the desired cytocompatibility against Human skin fibroblast (HFF-1 cells) with over 80% viability at the highest concentration and effective antibacterial activity against gram-positive and gram-negative bacteria (further improved by cross-linking with CDs and incorporating BEX), respectively. The incorporation of BEX and CDs in hydrogel films significantly enhanced the film's blood-clotting ability with negligible hemolysis due to blood clotting index and hemolysis tests.</p><p><strong>Conclusions: </strong>The findings of this study highlight the potential of biomaterial-based nano hydrogel film, composed of CS cross-linked with CDs and containing BEX, as a promising wound dressing with outstanding biocompatibility, minimal cytotoxicity, enhanced hemostatic efficacy, and strong antibacterial properties.</p>","PeriodicalId":15053,"journal":{"name":"Journal of Biological Engineering","volume":"19 1","pages":"31"},"PeriodicalIF":5.7,"publicationDate":"2025-04-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11987453/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144025969","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Development of a CRISPR-based cytosine base editor for restriction-modification system inactivation to enhance transformation efficiency in Vibrio Sp. dhg.","authors":"Yang Jun Shon, Dongyeop Baek, Su Bin Jin, Woo Jae Kim, Gyoo Yeol Jung, Hyun Gyu Lim","doi":"10.1186/s13036-025-00500-4","DOIUrl":"https://doi.org/10.1186/s13036-025-00500-4","url":null,"abstract":"<p><strong>Background: </strong>Vibrio sp. dhg is a fast-growing, alginate-utilizing, marine bacterium being developed as a platform host for macroalgae biorefinery. To maximize its potential in the production of various value-added products, there is a need to expand genetic engineering tools for versatile editing.</p><p><strong>Results: </strong>The CRISPR-based cytosine base editing (CBE) system was established in Vibrio sp. dhg, enabling C: G-to-T: A point mutations in multiple genomic loci. This CBE system displayed high editing efficiencies for single and multiple targets, reaching up to 100%. The CBE system efficiently introduced premature stop codons, inactivating seven genes encoding putative restriction enzymes of the restriction-modification system in two rounds. A resulting engineered strain displayed significantly enhanced transformation efficiency by up to 55.5-fold.</p><p><strong>Conclusions: </strong>Developing a highly efficient CBE system and improving transformation efficiency enable versatile genetic manipulation of Vibrio sp. dhg for diverse engineering in brown macroalgae bioconversion.</p>","PeriodicalId":15053,"journal":{"name":"Journal of Biological Engineering","volume":"19 1","pages":"30"},"PeriodicalIF":5.7,"publicationDate":"2025-04-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11984283/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144006702","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Termination sequence between an inducible promoter and ubiquitous chromatin opening element (UCOE) reduces gene expression leakage and silencing.","authors":"Tomoki Yanagi, Shean Fu Phen, Jonah Ayala, Deniz Ece Aydin, Susanna Jaramillo, David M Truong","doi":"10.1186/s13036-025-00499-8","DOIUrl":"https://doi.org/10.1186/s13036-025-00499-8","url":null,"abstract":"<p><strong>Background: </strong>Inducible gene expression circuits enable precise control over target gene activation and are widely used in direct reprogramming. However, their usability is often compromised by DNA methylation-induced silencing, especially in iPSCs. This deactivates genetic circuits in engineered iPSCs preventing them from being used for long-term scalable expansion of desired cell types. A2-ubiquitous chromatin opening elements (A2UCOE) have been recognized for their anti-silencing properties, but they have not been used in human iPSCs with inducible systems for direct reprogramming. This study investigates the role of A2UCOE in inducible systems and identifies strategies to eliminate associated gene leakage enabling long-term use of engineered human iPSCs.</p><p><strong>Results: </strong>We developed a compact all-in-one gene circuit - containing a doxycycline-inducible Tet-On system, 863 bp of A2UCOE, and FOXN1, a transcription factor critical for thymic epithelial cell (TEC) differentiation - easily deployed to new genomic sites. However, we observed significant FOXN1 gene leakage even without doxycycline, which is a novel limitation of A2UCOE. This leakage resulted in premature differentiation of iPSCs into TECs, limiting its continued use. To further investigate the relationship between A2UCOE and gene leakage, we generated A2UCOE fragments of varying lengths (1337 bp, 749 bp, and 547 bp) and found that all fragments, regardless of length, caused significant gene leakage. To solve this issue, we tested different spacer sequences between A2UCOE and the inducible promoter and found that the SV40 poly-A terminator fully eliminated FOXN1 leakage, and we show this effect is not due to AT- or GC-content. Unexpectedly, this architecture further enhanced anti-silencing effects > 60% providing prolonged stability for at least 30 days.</p><p><strong>Conclusions: </strong>This study reveals a novel limitation of A2UCOE in inducible systems, specifically its contribution to gene leakage, which compromise sensitive systems like direct reprogramming of iPSCs. The inclusion of an SV40 poly-A sequence provides a practical solution and genomic architecture to improve the functionality of A2UCOE-based circuits. It also suggests investigating how termination of transcription modulates gene silencing as a novel design parameter. These findings have significant implications for the design of robust gene circuits, particularly in applications involving iPSCs, regenerative medicine, and cell therapy.</p>","PeriodicalId":15053,"journal":{"name":"Journal of Biological Engineering","volume":"19 1","pages":"29"},"PeriodicalIF":5.7,"publicationDate":"2025-04-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11983960/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144006042","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}