New biotechnology最新文献_第10页

ReViTA: A novel in vitro transcription system to study gene regulation ReViTA：一种研究基因调控的新型体外转录系统

IF 5.4 2区生物学

New biotechnology Pub Date : 2023-09-25 DOI: 10.1016/j.nbt.2023.04.005

Alba Rubio-Canalejas , Lucas Pedraz , Eduard Torrents

{"title":"ReViTA: A novel in vitro transcription system to study gene regulation","authors":"Alba Rubio-Canalejas , Lucas Pedraz , Eduard Torrents","doi":"10.1016/j.nbt.2023.04.005","DOIUrl":"10.1016/j.nbt.2023.04.005","url":null,"abstract":"<div>ReViTA (Reverse in Vitro Transcription Assay) is a novel in vitro transcription-based method to study gene expression under the regulation of specific transcription factors. The ReViTA system uses a plasmid with a control sequence, the promoter region of the studied gene, the transcription factor of interest, and an RNA polymerase saturated with σ70. The main objective of this study was to evaluate the method; thus, as a proof of concept, two different transcription factors were used, a transcriptional inducer, AlgR, and a repressor, LexA, from Pseudomonas aeruginosa. After the promoters were incubated with the transcription factors, the plasmid was transcribed into RNA and reverse transcribed to cDNA. Gene expression was measured using qRT<img>PCR. Using the ReViTA plasmid, transcription induction of 55% was observed when AlgR protein was added and a 27% transcription reduction with the repressor LexA, compared with the samples without transcription factors. The results demonstrated the correct functioning of ReViTA as a novel method to study transcription factors and gene expression. Thus, ReViTA could be a rapid and accessible in vitro method to evaluate genes and regulators of various species.</div>","PeriodicalId":19190,"journal":{"name":"New biotechnology","volume":null,"pages":null},"PeriodicalIF":5.4,"publicationDate":"2023-09-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9647200","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

Purification of DNA oligonucleotides to improve hybridization chain reaction performance 纯化DNA寡核苷酸以提高杂交链反应性能

IF 5.4 2区生物学

New biotechnology Pub Date : 2023-09-25 DOI: 10.1016/j.nbt.2023.04.004

Mattias Leino, Ola Söderberg

引用次数: 0

Genetic code expansion in E. coli enables production of a functional ‘ready-to-click’ T cell receptor-specific scFv 大肠杆菌的遗传密码扩增使生产功能性“随时可点击”的T细胞受体特异性scFv成为可能

IF 5.4 2区生物学

New biotechnology Pub Date : 2023-09-25 DOI: 10.1016/j.nbt.2023.05.007

Rajeev Pasupuleti , Francesca Rosato , Dajana Kolanovic , Olga N. Makshakova , Winfried Römer , Birgit Wiltschi

{"title":"Genetic code expansion in E. coli enables production of a functional ‘ready-to-click’ T cell receptor-specific scFv","authors":"Rajeev Pasupuleti , Francesca Rosato , Dajana Kolanovic , Olga N. Makshakova , Winfried Römer , Birgit Wiltschi","doi":"10.1016/j.nbt.2023.05.007","DOIUrl":"10.1016/j.nbt.2023.05.007","url":null,"abstract":"<div>Antibody-based cancer therapies have been evolving at a rapid pace in the pharmaceutical market. Bispecific antibody-drug conjugates that engage immune cells to target and kill cancer cells with precision have inspired the development of immunotherapy. Miniaturized antibody fragments such as diabodies, nanobodies, or single-chain variable fragments (scFvs) hold great promise as antibody-drug conjugates as they specifically target tumor tissue and can penetrate it. Here, we optimized the soluble periplasmic expression of the scFv OKT3 comprising the variable VH and VL domains of the mouse anti-human CD3 antibody muromonab-CD3 (trade name Orthoclone OKT3) in E. coli. By an expansion of the genetic code, we site-specifically incorporated the reactive non-canonical amino acid Nε-((2-azidoethoxy)carbonyl)-L-lysine (AzK) into scFv OKT3 using an orthogonal pyrrolysyl-tRNA synthetase/tRNACUA pair. To confirm the AzK incorporation and to demonstrate the accessibility of the reactive azide group, we conjugated a fluorophore to scFv OKT3 AzK variants by copper-free strain-promoted alkyne-azide cycloaddition (‘click chemistry’). The scFv OKT3 wild type and the AzK variants bound T cells at nanomolar concentrations. In this study, a ‘ready-to-click’ scFv OKT3 was successfully developed for future applications, e.g. as controlled anti-T cell antibody-drug conjugate or bispecific T cell engager and for imaging immune T cell migration in cancers.</div>","PeriodicalId":19190,"journal":{"name":"New biotechnology","volume":null,"pages":null},"PeriodicalIF":5.4,"publicationDate":"2023-09-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9650272","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

An approach to rapid distributed manufacturing of broad spectrum anti-viral griffithsin using cell-free systems to mitigate pandemics 一种使用无细胞系统快速分布式生产广谱抗病毒Griffin的方法，以缓解流行病。

IF 5.4 2区生物学

New biotechnology Pub Date : 2023-09-25 DOI: 10.1016/j.nbt.2023.04.003

Shayan G. Borhani , Max Z. Levine , Lauren H. Krumpe , Jennifer Wilson , Curtis J. Henrich , Barry R. O’Keefe , Donald C. Lo , G. Sitta Sittampalam , Alexander G. Godfrey , R. Dwayne Lunsford , Venkata Mangalampalli , Dingyin Tao , Christopher A. LeClair , Aaron P. Thole , Douglas Frey , James Swartz , Govind Rao

{"title":"An approach to rapid distributed manufacturing of broad spectrum anti-viral griffithsin using cell-free systems to mitigate pandemics","authors":"Shayan G. Borhani , Max Z. Levine , Lauren H. Krumpe , Jennifer Wilson , Curtis J. Henrich , Barry R. O’Keefe , Donald C. Lo , G. Sitta Sittampalam , Alexander G. Godfrey , R. Dwayne Lunsford , Venkata Mangalampalli , Dingyin Tao , Christopher A. LeClair , Aaron P. Thole , Douglas Frey , James Swartz , Govind Rao","doi":"10.1016/j.nbt.2023.04.003","DOIUrl":"10.1016/j.nbt.2023.04.003","url":null,"abstract":"<div>This study describes the cell-free biomanufacturing of a broad-spectrum antiviral protein, griffithsin (GRFT) such that it can be produced in microgram quantities with consistent purity and potency in less than 24 h. We demonstrate GRFT production using two independent cell-free systems, one plant and one microbial. Griffithsin purity and quality were verified using standard regulatory metrics. Efficacy was demonstrated in vitro against SARS-CoV-2 and HIV-1 and was nearly identical to that of GRFT expressed in vivo. The proposed production process is efficient and can be readily scaled up and deployed wherever a viral pathogen might emerge. The current emergence of viral variants of SARS-CoV-2 has resulted in frequent updating of existing vaccines and loss of efficacy for front-line monoclonal antibody therapies. Proteins such as GRFT with its efficacious and broad virus neutralizing capability provide a compelling pandemic mitigation strategy to promptly suppress viral emergence at the source of an outbreak.</div>","PeriodicalId":19190,"journal":{"name":"New biotechnology","volume":null,"pages":null},"PeriodicalIF":5.4,"publicationDate":"2023-09-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10330340/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9759850","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: 3

A single-chain variable fragment selected against a conformational epitope of a recombinantly produced snake toxin using phage display 利用噬菌体展示筛选抗重组产生的蛇毒素构象表位的单链可变片段

IF 5.4 2区生物学

New biotechnology Pub Date : 2023-09-25 DOI: 10.1016/j.nbt.2023.04.002

Charlotte Rimbault , Pelle D. Knudsen , Anna Damsbo , Kim Boddum , Hanif Ali , Celeste M. Hackney , Lars Ellgaard , Markus-Frederik Bohn , Andreas H. Laustsen

{"title":"A single-chain variable fragment selected against a conformational epitope of a recombinantly produced snake toxin using phage display","authors":"Charlotte Rimbault , Pelle D. Knudsen , Anna Damsbo , Kim Boddum , Hanif Ali , Celeste M. Hackney , Lars Ellgaard , Markus-Frederik Bohn , Andreas H. Laustsen","doi":"10.1016/j.nbt.2023.04.002","DOIUrl":"10.1016/j.nbt.2023.04.002","url":null,"abstract":"<div>Phage display technology is a powerful tool for selecting monoclonal antibodies against a diverse set of antigens. Within toxinology, however, it remains challenging to generate monoclonal antibodies against many animal toxins, as they are difficult to obtain from venom. Recombinant toxins have been proposed as a solution to overcome this challenge, but so far, few have been used as antigens to generate neutralizing antibodies. Here, we describe the recombinant expression of α-cobratoxin in E. coli and its successful application as an antigen in a phage display selection campaign. From this campaign, an scFv (single-chain variable fragment) was isolated with similar binding affinity to a control scFv generated against the native toxin. The selected scFv recognizes a structural epitope, enabling it to inhibit the interaction between the acetylcholine receptor and the native toxin in vitro. This approach represents the first entirely in vitro antibody selection strategy for generating neutralizing monoclonal antibodies against a snake toxin.</div>","PeriodicalId":19190,"journal":{"name":"New biotechnology","volume":null,"pages":null},"PeriodicalIF":5.4,"publicationDate":"2023-09-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10004828","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: 2

Influence of feedstock source on the development of polyhydroxyalkanoates-producing mixed microbial cultures in continuously stirred tank reactors 原料来源对在连续搅拌釜式反应器中生产混合微生物培养物的聚羟基烷酸酯发展的影响

IF 5.4 2区生物学

New biotechnology Pub Date : 2023-09-25 DOI: 10.1016/j.nbt.2023.05.005

Elisa Clagnan, Fabrizio Adani

{"title":"Influence of feedstock source on the development of polyhydroxyalkanoates-producing mixed microbial cultures in continuously stirred tank reactors","authors":"Elisa Clagnan, Fabrizio Adani","doi":"10.1016/j.nbt.2023.05.005","DOIUrl":"10.1016/j.nbt.2023.05.005","url":null,"abstract":"<div>Polyhydroxyalkanoates (PHAs) are the new frontier of bioplastic production; however, research is needed to develop and characterise efficient mixed microbial communities (MMCs) for their application with a multi-feedstock approach. Here, the performance and composition of six MMCs developed from the same inoculum on different feedstocks were investigated through Illumina sequencing to understand community development and identify possible redundancies in terms of genera and PHA metabolism. High PHA production efficiencies (>80% mg CODPHA mg-1 CODOA-consumed) were seen across all samples, but differences in the organic acids (OAs) composition led to different ratios of the monomers poly(3-hydroxybutyrate) (3HB) to poly(3-hydroxyvalerate) (3HV). Communities differed across all feedstocks, with enrichments in specific PHA-producing genera, but analysis of potential enzymatic activity identified a certain degree of functional redundancy, possibly leading to the general high efficiency seen in PHA production from all feedstocks. Leading PHAs producers across all feedstocks were identified in genera such as Thauera, Leadbetterella, Neomegalonema and Amaricoccus.</div>","PeriodicalId":19190,"journal":{"name":"New biotechnology","volume":null,"pages":null},"PeriodicalIF":5.4,"publicationDate":"2023-09-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9655740","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

Domain shifts in dermoscopic skin cancer datasets: Evaluation of essential limitations for clinical translation 皮肤镜下皮肤癌数据集的领域转移:评估临床翻译的基本限制

IF 5.4 2区生物学

New biotechnology Pub Date : 2023-09-25 DOI: 10.1016/j.nbt.2023.04.006

Katharina Fogelberg , Sireesha Chamarthi , Roman C. Maron , Julia Niebling , Titus J. Brinker

{"title":"Domain shifts in dermoscopic skin cancer datasets: Evaluation of essential limitations for clinical translation","authors":"Katharina Fogelberg , Sireesha Chamarthi , Roman C. Maron , Julia Niebling , Titus J. Brinker","doi":"10.1016/j.nbt.2023.04.006","DOIUrl":"10.1016/j.nbt.2023.04.006","url":null,"abstract":"<div>The limited ability of Convolutional Neural Networks to generalize to images from previously unseen domains is a major limitation, in particular, for safety-critical clinical tasks such as dermoscopic skin cancer classification. In order to translate CNN-based applications into the clinic, it is essential that they are able to adapt to domain shifts. Such new conditions can arise through the use of different image acquisition systems or varying lighting conditions. In dermoscopy, shifts can also occur as a change in patient age or occurrence of rare lesion localizations (e.g. palms). These are not prominently represented in most training datasets and can therefore lead to a decrease in performance. In order to verify the generalizability of classification models in real world clinical settings it is crucial to have access to data which mimics such domain shifts. To our knowledge no dermoscopic image dataset exists where such domain shifts are properly described and quantified. We therefore grouped publicly available images from ISIC archive based on their metadata (e.g. acquisition location, lesion localization, patient age) to generate meaningful domains. To verify that these domains are in fact distinct, we used multiple quantification measures to estimate the presence and intensity of domain shifts. Additionally, we analyzed the performance on these domains with and without an unsupervised domain adaptation technique. We observed that in most of our grouped domains, domain shifts in fact exist. Based on our results, we believe these datasets to be helpful for testing the generalization capabilities of dermoscopic skin cancer classifiers.</div>","PeriodicalId":19190,"journal":{"name":"New biotechnology","volume":null,"pages":null},"PeriodicalIF":5.4,"publicationDate":"2023-09-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10006368","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 recombinant strain of Komagataeibacter xylinus ATCC 23770 for production of bacterial cellulose from mannose-rich resources 利用富含甘露糖的资源生产细菌纤维素的木霉菌ATCC 23770重组菌株

IF 5.4 2区生物学

New biotechnology Pub Date : 2023-09-25 DOI: 10.1016/j.nbt.2023.05.002

Fan Yang , Zhangjun Cao , Can Li , Lin Chen , Guochao Wu , Xingping Zhou , Feng F. Hong

{"title":"A recombinant strain of Komagataeibacter xylinus ATCC 23770 for production of bacterial cellulose from mannose-rich resources","authors":"Fan Yang , Zhangjun Cao , Can Li , Lin Chen , Guochao Wu , Xingping Zhou , Feng F. Hong","doi":"10.1016/j.nbt.2023.05.002","DOIUrl":"10.1016/j.nbt.2023.05.002","url":null,"abstract":"<div>The development of bacterial cellulose (BC) industrialization has been seriously affected by its production. Mannose/mannan is an essential component in many biomass resources, but Komagataeibacter xylinus uses mannose in an ineffective way, resulting in waste. The aim of this study was to construct recombinant bacteria to use mannose-rich biomass efficiently as an alternative and inexpensive carbon source in place of the more commonly used glucose. This strategy aimed at modification of the mannose catabolic pathway via genetic engineering of K. xylinus ATCC 23770 strain through expression of mannose kinase and phosphomannose isomerase genes from the Escherichia coli K-12 strain. Recombinant and wild-type strains were cultured under conditions of glucose and mannose respectively as sole carbon sources. The fermentation process and physicochemical properties of BC were investigated in detail in the strains cultured in mannose media. The comparison showed that with mannose as the sole carbon source, the BC yield from the recombinant strain increased by 84%, and its tensile strength and elongation were increased 1.7 fold, while Young's modulus was increased 1.3 fold. The results demonstrated a successful improvement in BC yield and properties on mannose-based medium compared with the wild-type strain. Thus, the strategy of modifying the mannose catabolic pathway of K. xylinus is feasible and has significant potential in reducing the production costs for industrial production of BC from mannose-rich biomass.</div>","PeriodicalId":19190,"journal":{"name":"New biotechnology","volume":null,"pages":null},"PeriodicalIF":5.4,"publicationDate":"2023-09-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9642281","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: 2

Valorization of bioethanol by-products to produce unspecific peroxygenase with Agrocybe aegerita: Technological and proteomic perspectives 生物乙醇副产物的增值利用绿草结霉生产非特异性过氧酶:技术和蛋白质组学观点

IF 5.4 2区生物学

New biotechnology Pub Date : 2023-09-25 DOI: 10.1016/j.nbt.2023.05.001

Sandra González-Rodríguez, Alba Trueba-Santiso, Thelmo A. Lu-Chau, María Teresa Moreira, Gemma Eibes

引用次数: 0

A step closer to continuous buffer preparation from solids: Predicting powder compaction and how to prevent it 从固体中连续制备缓冲剂的一步:预测粉末压实和如何防止它

IF 5.4 2区生物学

New biotechnology Pub Date : 2023-09-25 DOI: 10.1016/j.nbt.2023.05.003

D. Komuczki , N. Hesse , J. Schmidt , P. Satzer

{"title":"A step closer to continuous buffer preparation from solids: Predicting powder compaction and how to prevent it","authors":"D. Komuczki , N. Hesse , J. Schmidt , P. Satzer","doi":"10.1016/j.nbt.2023.05.003","DOIUrl":"10.1016/j.nbt.2023.05.003","url":null,"abstract":"<div>The preparation of buffer solutions used in the biopharmaceutical industry is typically performed manually by the addition of one or multiple buffering reagents to water. Recently, the adaptation of powder feeders for continuous solid feeding was demonstrated for continuous buffer preparation. However, the intrinsic characteristics of powders can change the stability of the process, due to the hygroscopic nature of some substances and humidity-induced caking and compaction behavior, but there is no simple and easy methodology available for predicting this behavior for buffer species. To predict which buffering reagents are suitable without special precautions and investigate their behavior, force displacement measurements were conducted with a customized rheometer over 18 h. While most of the eight investigated buffering reagents indicated uniform compaction, especially sodium acetate and dipotassium hydrogen phosphate (K2HPO4) showed a significant increase in yield stress after 2 h. Experiments conducted with a 3D printed miniaturized screw conveyor confirmed the increased yield stress measurements by visible compaction and failure of the feeding. By taking additional precautions and adjusting the design of the hopper, we demonstrated a highly linear profile of all buffering reagents over a duration of 12 and 24 h. We showed that force displacement measurements accurately predict the behavior of buffer components in continuous feeding devices for continuous buffer preparation and are a valuable tool to identify buffer components that need special precautions. Stable, precise feeding of all tested buffer components was demonstrated, highlighting the importance of identifying buffers that need a specialized setup with a rapid methodology.</div>","PeriodicalId":19190,"journal":{"name":"New biotechnology","volume":null,"pages":null},"PeriodicalIF":5.4,"publicationDate":"2023-09-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9675445","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