New biotechnology最新文献_第5页

Stepwise activation of gene copies results in higher final titers of subclones compared to immediate integration of the full set of active copies 与立即整合全套活性拷贝相比，基因拷贝的逐步激活导致亚克隆的最终滴度更高

IF 4.5 2区生物学

New biotechnology Pub Date : 2025-04-24 DOI: 10.1016/j.nbt.2025.04.011

Víctor Jiménez Lancho , Peter Eisenhut , Gerald Klanert , Daniel Ivansson , Andreas Jonsson , Ann Lövgren , Nicole Borth

{"title":"Stepwise activation of gene copies results in higher final titers of subclones compared to immediate integration of the full set of active copies","authors":"Víctor Jiménez Lancho , Peter Eisenhut , Gerald Klanert , Daniel Ivansson , Andreas Jonsson , Ann Lövgren , Nicole Borth","doi":"10.1016/j.nbt.2025.04.011","DOIUrl":"10.1016/j.nbt.2025.04.011","url":null,"abstract":"<div><div>The increasing demand for production of therapeutic proteins has encouraged both industrial and academic institutions to pursue the development of mammalian expression platforms with high productivities. While protocols for rapid and efficient integration of multiple transgene copies into the genome are available, they require substantial time and resources for screening numerous clones. A contributing factor is the tendency of high producers to disappear from the selected mini-pools due to the stress caused by high productivity without adequate time for adaptation of cellular capacities. Here, we have developed a strategy to stably activate individual copies within an initially repressed multicopy coding cassette harboring 2 GFP-Fc and 2 BFP-Fc genes, each fused to an Fc region for secretion. This toolbox enables gene activation via CRISPR/Cas9-mediated deletion of the repressor elements. Subsequently, producers can be sorted based on increased GFP or BFP fluorescence and assessed by measuring the secreted total Fc protein. We demonstrate that the stepwise activation of initially repressed genes outperforms a control cell line with the same number of genes active from the outset, as evidenced by higher fluorescence signals from GFP and BFP, increased mRNA levels for BFP, GFP, and Fc genes, and enhanced titer of secreted Fc fusion protein. This study demonstrates the ability of cells to adapt to new challenges by modulating both gene expression patterns and channeling of resources to accommodate high production loads.</div></div>","PeriodicalId":19190,"journal":{"name":"New biotechnology","volume":"88 ","pages":"Pages 89-99"},"PeriodicalIF":4.5,"publicationDate":"2025-04-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143890889","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Open-source antibodies as a path to enhanced research reproducibility and transparency 开源抗体是提高研究可重复性和透明度的途径

IF 4.5 2区生物学

New biotechnology Pub Date : 2025-04-17 DOI: 10.1016/j.nbt.2025.04.004

Meghan Rego , Douglas W. Houston , Melina Fan , Karl D. Murray , James S. Trimmer

{"title":"Open-source antibodies as a path to enhanced research reproducibility and transparency","authors":"Meghan Rego , Douglas W. Houston , Melina Fan , Karl D. Murray , James S. Trimmer","doi":"10.1016/j.nbt.2025.04.004","DOIUrl":"10.1016/j.nbt.2025.04.004","url":null,"abstract":"<div><div>Antibodies are important tools with diverse uses in biomedical research. However, open access to reliable sources of well-characterized antibodies with unambiguous molecular identities remains an obstacle to research transparency and reproducibility. We propose here a community shift towards open-source antibodies, analogous to open-source computer software. The tenets of such antibodies are that 1) they are available to researchers in a ready to use form, 2) the renewable source of the antibody (<em>e.g.</em>, hybridoma cells or plasmid) is also widely available ensuring reproducible and cost-effective access to the same antibody, and 3) the antibody sequence is publicly available. With these criteria met, the antibody can be widely used with the transparent assurance associated with a molecularly defined reagent, and the code can be edited to generate antibody variants to meet researchers’ specific needs. We (the UC Davis/NIH NeuroMab Facility, the Development Studies Hybridoma Bank, and Addgene) have established a consortium to provide open-source access to a large collection of well characterized antibodies. As open-source software has benefitted both users and developers, we suggest open-source antibodies will have a similar positive impact on antibody based biomedical research. We encourage funding agencies to support initiatives to expand access to open-source antibody resources, and researchers to both utilize and to contribute to them, with a goal of enabling more reliable and cost-effective pursuit of research.</div></div>","PeriodicalId":19190,"journal":{"name":"New biotechnology","volume":"87 ","pages":"Pages 121-129"},"PeriodicalIF":4.5,"publicationDate":"2025-04-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143870713","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Site-directed antibodies targeting driver mutations of the KRAS protein 靶向KRAS蛋白驱动突变的定点抗体

IF 4.5 2区生物学

New biotechnology Pub Date : 2025-04-17 DOI: 10.1016/j.nbt.2025.04.003

Xiaofeng Li , Qiana Mendez , Cassandra Chapados , Felicity Acca , Holly Driscoll , Jason Oliveira , Jun Liu , Kezzia Jones , Mary Ferguson , Ryan L. Wallace , Sergei Bibikov , Troy Lionberger , Kevin J. Harvey , Michael P. Weiner , Greg Mirando

{"title":"Site-directed antibodies targeting driver mutations of the KRAS protein","authors":"Xiaofeng Li , Qiana Mendez , Cassandra Chapados , Felicity Acca , Holly Driscoll , Jason Oliveira , Jun Liu , Kezzia Jones , Mary Ferguson , Ryan L. Wallace , Sergei Bibikov , Troy Lionberger , Kevin J. Harvey , Michael P. Weiner , Greg Mirando","doi":"10.1016/j.nbt.2025.04.003","DOIUrl":"10.1016/j.nbt.2025.04.003","url":null,"abstract":"<div><div>Kirsten rat sarcoma viral oncogene homolog (KRAS) is the most mutated oncogene in human cancers, found in approximately 30 % of tumors. These mutations primarily consist of single-base missense alterations in codon G12. While extensive efforts have focused on developing allele-specific inhibitors for KRAS mutations, mutation-specific antibodies (Abs) remain largely unexplored, with only a few research-use-only catalog Abs available. In this study, we employed the proprietary Epivolve technology to develop site-directed monoclonal Abs (mAbs) that target KRAS oncogenic driver mutation KRAS G12D. These site-directed mAbs demonstrate high binding affinity, with equilibrium dissociation constants (K<sub>D</sub>) in the nanomolar range, showing over 1,000-fold greater affinity for KRAS G12D compared to wild-type KRAS. Western blot analyses using both purified KRAS protein variants and tumor cell lines harboring G12D mutations confirmed the high specificity of these mAbs. Furthermore, immunocytochemistry analysis revealed co-localization of the site-directed mAbs with endogenously expressed KRAS in cancer cells bearing G12D mutations. The validated high affinity and specificity of these site-directed mAbs highlight their potential for diagnostic applications and therapeutic development targeting KRAS driver mutations.</div></div>","PeriodicalId":19190,"journal":{"name":"New biotechnology","volume":"87 ","pages":"Pages 112-120"},"PeriodicalIF":4.5,"publicationDate":"2025-04-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143870714","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Systematic identification of genomic hotspots for high-yield protein production in CHO cells CHO细胞高产蛋白生产基因组热点的系统鉴定

IF 4.5 2区生物学

New biotechnology Pub Date : 2025-04-12 DOI: 10.1016/j.nbt.2025.04.006

Minouk Lee , Sung-Hyuk Han , Dongseok Kim , Seongtae Yun , Jinho Yeom , Minji Kyeong , Seo-Young Park , Dong-Yup Lee

{"title":"Systematic identification of genomic hotspots for high-yield protein production in CHO cells","authors":"Minouk Lee , Sung-Hyuk Han , Dongseok Kim , Seongtae Yun , Jinho Yeom , Minji Kyeong , Seo-Young Park , Dong-Yup Lee","doi":"10.1016/j.nbt.2025.04.006","DOIUrl":"10.1016/j.nbt.2025.04.006","url":null,"abstract":"<div><div>The efficient and stable production of therapeutic proteins in Chinese hamster ovary (CHO) cells hinges on robust cell line development (CLD). Traditional methods relying on random transgene integration often result in clonal variability, requiring extensive and resource-intensive screening. To address this limitation, we established a systematic, multiomics-driven framework that integrates 202 RNA-sequencing datasets and whole-genome sequencing data to identify genomic “hotspot” loci for precise and high-yield transgene integration. From an initial pool of 20 candidate loci, 5 top-performing hotspots were validated using site-specific integration in CHO-DG44 cells via the CRISPR/Cas9 system with Recombinase-mediated cassette exchange (RMCE). These genomic hotspots achieved 2.2- to 15.0-fold higher relative specific productivity compared to previously known controls (<em>Fer1L4</em> and <em>Locus1</em> sites), across multiple therapeutic proteins, including a lysosomal storage disorder-related enzyme and an Immunoglobulin G (IgG)-related monoclonal antibody (mAb) expression. This study offers a transformative approach to CLD, achieving significant improvements in productivity, genomic stability, and efficiency, as well as paving the way for enhanced biopharmaceutical manufacturing.</div></div>","PeriodicalId":19190,"journal":{"name":"New biotechnology","volume":"88 ","pages":"Pages 61-72"},"PeriodicalIF":4.5,"publicationDate":"2025-04-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143829313","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Valorization of waste cooking oil for bioproduction of industrially-relevant metabolites in Ashbya gossypii 废食用油在棉叶Ashbya gossypii中用于工业相关代谢物生物生产的价值评价

IF 4.5 2区生物学

New biotechnology Pub Date : 2025-04-07 DOI: 10.1016/j.nbt.2025.04.002

Javier Martín-González , Javier-Fernando Montero-Bullón , Gloria Muñoz-Fernández , Rubén M. Buey , Alberto Jiménez

{"title":"Valorization of waste cooking oil for bioproduction of industrially-relevant metabolites in Ashbya gossypii","authors":"Javier Martín-González , Javier-Fernando Montero-Bullón , Gloria Muñoz-Fernández , Rubén M. Buey , Alberto Jiménez","doi":"10.1016/j.nbt.2025.04.002","DOIUrl":"10.1016/j.nbt.2025.04.002","url":null,"abstract":"<div><div>Waste cooking oil (WCO) is a byproduct of culinary processes, which undergoes degradation due to high temperatures during frying and cooking. Beyond its detrimental effects on health, including potential carcinogenic effects, WCO poses a significant environmental threat, emphasizing the need for urgent recycling efforts. Valorization of WCO as a carbon source for microbial fermentations emerges as a feasible alternative in a bioeconomy context. The aim of the present work is to explore the ability of <em>Ashbya gossypii</em>, a natural overproducer of riboflavin that is currently used in the industrial production of the vitamin, to exploit WCO for the production of industrially relevant metabolites such as riboflavin, folates, biolipids and monoterpenes. Our results demonstrate that WCO is an effective carbon source for <em>A. gossypii</em> bioproduction of riboflavin, folates and biolipids, reaching among the highest titers described so far in flask fermentation: riboflavin titer (312.5 mg/L) increased 4.8-fold compared to glucose-based medium; folate production reached 7.6 mg/L; and the intracellular lipids were above 80 % of the cell dry weight. In contrast, the production of the monoterpenes limonene and sabinene was not improved with the utilization of WCO. Taken together, our results present a proof-of-principle for the implementation of a novel bioprocess for the valorization of WCO using the industrial fungus <em>A. gossypii</em>.</div></div>","PeriodicalId":19190,"journal":{"name":"New biotechnology","volume":"88 ","pages":"Pages 32-38"},"PeriodicalIF":4.5,"publicationDate":"2025-04-07","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143791937","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Screening assay for polyester hydrolyzing microorganisms using fluorescence-labeled poly(butylene adipate) 利用荧光标记聚己二酸丁二醇酯筛选聚酯水解微生物的检测方法。

IF 4.5 2区生物学

New biotechnology Pub Date : 2025-04-05 DOI: 10.1016/j.nbt.2025.03.007

Bettina Semler , Karin Binder , Doris Ribitsch , Alessandro Pellis , Georg M. Guebitz

{"title":"Screening assay for polyester hydrolyzing microorganisms using fluorescence-labeled poly(butylene adipate)","authors":"Bettina Semler , Karin Binder , Doris Ribitsch , Alessandro Pellis , Georg M. Guebitz","doi":"10.1016/j.nbt.2025.03.007","DOIUrl":"10.1016/j.nbt.2025.03.007","url":null,"abstract":"<div><div>Despite recent advances, there is still a demand for more efficient enzymes hydrolyzing synthetic polymers. Automated high throughput screening strategies of microorganisms from different environments could yield novel enzymes but require specific methods for detection of polymer hydrolysis in complex matrices. Here, 5-carboxy-fluorescein (5-FAM) was covalently coupled to poly(butylene adipate) (PBA) and blended at 1 %, 5 % and 10 % w/w concentrations with non-labeled PBA. Hydrolysis of PBA by the Thc_Cut1 cutinase from <em>Thermobifida cellulosilytica</em> was confirmed via quantification of the released monomers 1,4-butanediol and adipic acid, weight loss and FTIR analysis. Upon incubation with Thc_Cut1, hydrolysis of all three fluorescent labeled PBA blends lead to a clear fluorescence increase of up to 4000 RFU while no signal change was detected for the blank and for heat-inactivated enzyme (signal below 500 RFU). In a next step, as a model organism <em>Pichia pastoris</em> expressing the identical cutinase was cultivated in the presences of labeled PBA. Despite the complex matrix, a fluorescence increase of up to 500 RFU was observed for <em>P. pastoris</em> expressing the enzyme while no significant signal change was seen for the control strain (lacking Thc_Cut1 expression). Likewise, extracellular enzymes from the fungi <em>Fusarium solani</em> and <em>Alternaria alternata</em> hydrolyzed labeled PBA leading to fluorescence increases of 1328 and 1187 RFU. This indicates that 5-FAM covalently coupled to polymers could be used for development of simple and high throughput screening platforms to identify polymer decomposing microorganisms and enzymes.</div></div>","PeriodicalId":19190,"journal":{"name":"New biotechnology","volume":"88 ","pages":"Pages 39-45"},"PeriodicalIF":4.5,"publicationDate":"2025-04-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143803799","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

A distinct autofluorescence distribution pattern marks enzymatic deconstruction of plant cell wall 一种独特的自身荧光分布模式标志着酶解植物细胞壁。

IF 4.5 2区生物学

New biotechnology Pub Date : 2025-04-05 DOI: 10.1016/j.nbt.2025.04.001

Solmaz Hossein Khani , Khadidja Ould Amer , Noah Remy , Berangère Lebas , Anouck Habrant , Ali Faraj , Grégoire Malandain , Gabriel Paës , Yassin Refahi

{"title":"A distinct autofluorescence distribution pattern marks enzymatic deconstruction of plant cell wall","authors":"Solmaz Hossein Khani , Khadidja Ould Amer , Noah Remy , Berangère Lebas , Anouck Habrant , Ali Faraj , Grégoire Malandain , Gabriel Paës , Yassin Refahi","doi":"10.1016/j.nbt.2025.04.001","DOIUrl":"10.1016/j.nbt.2025.04.001","url":null,"abstract":"<div><div>Achieving an economically viable transformation of plant cell walls into bioproducts requires a comprehensive understanding of enzymatic deconstruction. Microscale quantitative analysis offers a relevant approach to enhance our understanding of cell wall hydrolysis, but becomes challenging under high deconstruction conditions. This study comprehensively addresses the challenges of quantifying the impact of extensive enzymatic deconstruction on plant cell wall at microscale. Investigation of highly deconstructed spruce wood provided spatial profiles of cell walls during hydrolysis with remarkable precision. A distinct cell wall autofluorescence distribution pattern marking enzymatic hydrolysis along with an asynchronous impact of hydrolysis on cell wall structure, with cell wall volume reduction preceding cell wall accessible surface area decrease, were revealed. This study provides novel insights into enzymatic deconstruction of cell wall at under-investigated cell scale, and a robust computational pipeline applicable to diverse biomass species and pretreatment types for assessing hydrolysis impact and efficiency.</div></div>","PeriodicalId":19190,"journal":{"name":"New biotechnology","volume":"88 ","pages":"Pages 46-60"},"PeriodicalIF":4.5,"publicationDate":"2025-04-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143803784","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Inspired by nature: using plant virus particles in biomedicine and nanotechnology 灵感来自大自然：在生物医学和纳米技术中使用植物病毒颗粒

IF 4.5 2区生物学

New biotechnology Pub Date : 2025-03-25 DOI: 10.1016/j.nbt.2024.08.067

Lomonossoff G

引用次数: 0

Bridging Biotech: Revyve's Leap from Laboratory to Large-Scale Production 弥合生物技术：Revyve从实验室到大规模生产的飞跃

IF 4.5 2区生物学

New biotechnology Pub Date : 2025-03-25 DOI: 10.1016/j.nbt.2024.08.056

Van Den Berg C

引用次数: 0

EU-SAGE (European Sustainable Agriculture through Genome Editing): the role of scientists in the new genomic techniques regulation in Europe EU-SAGE（通过基因组编辑的欧洲可持续农业）：科学家在欧洲新基因组技术监管中的作用