BioTechniques最新文献_第2页

Label-free DNA-sensor modelling based on magnetic induction spectroscopy. 基于磁感应光谱的无标记dna传感器建模。

IF 2.5 4区工程技术

BioTechniques Pub Date : 2026-01-01 Epub Date: 2026-03-27 DOI: 10.1080/07366205.2026.2647885

César A González-Díaz, Boris Rubinsky, Simo A Mäkiharju, Virginia Sánchez-Monroy, Franck M André, Luis M Mir

引用次数: 0

Single-cell RNA-sequencing of air-liquid interface cultures: a practical approach. 气液界面培养的单细胞rna测序：一种实用的方法。

IF 2.5 4区工程技术

BioTechniques Pub Date : 2026-01-01 Epub Date: 2026-03-28 DOI: 10.1080/07366205.2026.2621052

Firoozeh Vosoogh-Gerayeli, Carolyn J Wang, Chen X Yang, Gurpreet S Singhera, Edward Z Li, Xuan Li, Julia S W Yang, Chung Y Cheung, Janice M Leung, Don D Sin

引用次数: 0

Optimized Southern blotting for enhanced and precise detection of transgenes in CHO cells from transposon-based expression systems. 优化的Southern印迹法用于从转座子表达系统中增强和精确检测CHO细胞中的转基因。

IF 2.5 4区工程技术

BioTechniques Pub Date : 2026-01-01 Epub Date: 2026-01-29 DOI: 10.1080/07366205.2026.2621051

Hyo-Young Jeong, Caitlyn Devine, Bor-Ruei Lin, Zhenqiu Huang, Guanghua Li, Lin Zhang

{"title":"Optimized Southern blotting for enhanced and precise detection of transgenes in CHO cells from transposon-based expression systems.","authors":"Hyo-Young Jeong, Caitlyn Devine, Bor-Ruei Lin, Zhenqiu Huang, Guanghua Li, Lin Zhang","doi":"10.1080/07366205.2026.2621051","DOIUrl":"10.1080/07366205.2026.2621051","url":null,"abstract":"The genetic stability of recombinant CHO cell lines producing therapeutic proteins is critical for ensuring consistent quality in biopharma-ceutical products. Southern blotting remains the gold standard for evaluating transgene integrity and stability in these cell lines. In the biopharmaceutical industry, transposon-based expression systems are widely utilized to generate highly productive and genetically stable CHO cell lines. However, evaluating transgene integration sites and integrity in such cell lines is challenging with standard Southern blotting protocols. This difficulty arises because transposon-mediated transfection often results in multiple independent integration sites in the host genome, each typically harboring a single transgene copy. Upon restriction enzyme digestion, similar-sized DNA fragments are generated, reducing resolution and complicating the separation and detection of the transgenes using standard blotting protocols. Here, we present a modified Southern blotting protocol that significantly improves the resolution of integration banding patterns by refining key steps, including purification of digested DNA prior to electrophoresis and an enhanced DNA transfer method. This protocol was successfully applied to analyze multiple transposon-derived CHO cell lines with high transgene copy numbers, enabling more precise and efficient detection of transgene integration.","PeriodicalId":8945,"journal":{"name":"BioTechniques","volume":" ","pages":"1-10"},"PeriodicalIF":2.5,"publicationDate":"2026-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"146084041","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Quantitative analysis of oligonucleotide delivery into isolated mitochondria: a proof-of-concept method. 寡核苷酸递送到分离线粒体的定量分析：一种概念验证方法。

IF 2.5 4区工程技术

BioTechniques Pub Date : 2026-01-01 Epub Date: 2026-03-26 DOI: 10.1080/07366205.2026.2635461

Joshua McHale, Veronica Bazzani, Abdechakour Elkihel, Jing Wu, Ling Peng, Carlo Vascotto

{"title":"Quantitative analysis of oligonucleotide delivery into isolated mitochondria: a proof-of-concept method.","authors":"Joshua McHale, Veronica Bazzani, Abdechakour Elkihel, Jing Wu, Ling Peng, Carlo Vascotto","doi":"10.1080/07366205.2026.2635461","DOIUrl":"https://doi.org/10.1080/07366205.2026.2635461","url":null,"abstract":"Mitochondria, with their own DNA, Represent a potential target for nucleic acid-based precision therapies. However, effective delivery of therapeutic oligonucleotides remains challenging due to the dual mitochondrial membranes and the localization of mitochondrial DNA within nucleoid complexes in the matrix. To understand the delivery process and assess the delivery efficiency of potential vectors, such as dendrimers, it is essential to effectively quantify the oligonucleotides that are successfully delivered to and remain within mitochondria. Currently, there are only limited yet inconvenient methods available for this purpose. Here, we describe a method for quantifying the delivery of fluorescent oligonucleotide cargos in isolated mitochondria using a microfiltration apparatus for reliable fluorescent analysis. By working within a range of dilutions, we are able to safeguard the concentration limits. The quantification protocol also enables the visualization of specific localization within mitochondria, allowing for the determination of whether delivery can occur across both membranes. This is particularly useful, as it offers a key insight into improving vectors as they must deliver the cargoes within the mitochondrial matrix. We validate this method in this proof-of-concept study, providing biological data to assess the difference between two amphiphilic dendrimer vectors for oligonucleotide delivery in mitochondria.","PeriodicalId":8945,"journal":{"name":"BioTechniques","volume":"78 1-12","pages":"1-11"},"PeriodicalIF":2.5,"publicationDate":"2026-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147509486","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Evaluation and verification of cfDNA extraction methods. cfDNA提取方法的评价与验证。

IF 2.5 4区工程技术

BioTechniques Pub Date : 2026-01-01 Epub Date: 2026-05-04 DOI: 10.1080/07366205.2026.2665280

Cecilie Mondrup Jacobsen, Luisa Matos do Canto, Estrid Høgdall, Rikke Fredslund Andersen

引用次数: 0

Comparative performance of high-throughput qPCR, qPCR, and digital PCR for antibiotic resistance gene monitoring in tropical water systems. 高通量qPCR、qPCR和数字PCR在热带水系统抗生素耐药基因监测中的比较性能

IF 2.5 4区工程技术

BioTechniques Pub Date : 2026-01-01 Epub Date: 2026-04-22 DOI: 10.1080/07366205.2026.2649861

Thitima Srathongneam, Phongsawat Paisantham, Andrew C Singer, Rojana Sukchawalit, Skorn Mongkolsuk, Kwanrawee Sirikanchana

{"title":"Comparative performance of high-throughput qPCR, qPCR, and digital PCR for antibiotic resistance gene monitoring in tropical water systems.","authors":"Thitima Srathongneam, Phongsawat Paisantham, Andrew C Singer, Rojana Sukchawalit, Skorn Mongkolsuk, Kwanrawee Sirikanchana","doi":"10.1080/07366205.2026.2649861","DOIUrl":"10.1080/07366205.2026.2649861","url":null,"abstract":"Antibiotic resistance genes (ARGs) are important contaminants in water systems, and their detection depends strongly on methodological sensitivity. This study compared three molecular platforms, high-throughput quantitative polymerase chain reaction (HT-qPCR), hydrolysis probe-based qPCR, and droplet digital PCR (ddPCR), for detecting ARGs in wastewater, river water, and seawater in Thailand. HT-qPCR enabled broad resistome profiling, detecting 325-336 ARGs out of 373 targets (87.1-90.1%), with aminoglycoside, beta-lactam, macrolide-lincosamide-streptogramin B, sulfonamide, mobile genetic element, and integron genes most prevalent. qPCR quantified selected ARGs using standard curves constructed from plasmid standards whose absolute copy numbers were calibrated by ddPCR, yielding high efficiency and strong linearity. ddPCR detected the same target genes as qPCR with comparable concentration estimates and additionally identified blaIND, which was not observed by HT-qPCR or qPCR. Quantifications from qPCR and ddPCR showed high rank-based concordance across matrices. Combining HT-qPCR with qPCR improved coverage by about 1% relative to HT-qPCR alone, while HT-qPCR with ddPCR offered nearly identical values. In conclusion, HT-qPCR was most effective for comprehensive profiling, qPCR ensured reliable quantification, and ddPCR enabled detection of rare targets, supporting a tiered and cost-effective framework for smart ARG monitoring in tropical aquatic environments.","PeriodicalId":8945,"journal":{"name":"BioTechniques","volume":"78 1-12","pages":"158-169"},"PeriodicalIF":2.5,"publicationDate":"2026-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147760898","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

High-resolution molecular typing of vancomycin-resistant Enterococcus faecium from Romania and Bavaria: combining enhanced DNA microarray and next generation sequencing. 罗马尼亚和巴伐利亚耐万古霉素屎肠球菌的高分辨率分子分型：结合增强DNA微阵列和下一代测序。

IF 2.5 4区工程技术

BioTechniques Pub Date : 2026-01-01 Epub Date: 2026-02-02 DOI: 10.1080/07366205.2026.2621054

Ibukun Elizabeth Osadare, Abdinasir Abdilahi, Elke Müller, Mara Lohde, Celia Diezel, Maximilian Collatz, Sascha Braun, Bärbel Kieninger, Anja Eichner, Wulf Schneider-Brachert, Thomas Wellhöfer, Katrin Frankenfeld, Olivia Dorneanu, Stefan Monecke, Ralf Ehricht

{"title":"High-resolution molecular typing of vancomycin-resistant Enterococcus faecium from Romania and Bavaria: combining enhanced DNA microarray and next generation sequencing.","authors":"Ibukun Elizabeth Osadare, Abdinasir Abdilahi, Elke Müller, Mara Lohde, Celia Diezel, Maximilian Collatz, Sascha Braun, Bärbel Kieninger, Anja Eichner, Wulf Schneider-Brachert, Thomas Wellhöfer, Katrin Frankenfeld, Olivia Dorneanu, Stefan Monecke, Ralf Ehricht","doi":"10.1080/07366205.2026.2621054","DOIUrl":"https://doi.org/10.1080/07366205.2026.2621054","url":null,"abstract":"Antimicrobial resistance poses a significant challenge for infection control, requiring the development of accurate and high-throughput diagnostic techniques. We expanded and optimized an existing DNA microarray platform for the molecular characterization of vancomycin-resistant Enterococcus (VRE) by incorporating resistance, virulence, species-specific, and typing markers. The enhanced microarray allows for the simultaneous analysis of up to 96 strains, providing detailed genetic profiles of clinical isolates. VRE strains from Romania and Bavaria, Germany, were analyzed, and the results were compared to those obtained using traditional typing methods, such as multilocus sequence typing (MLST). Next-generation sequencing (NGS) was used in parallel to validate the microarray findings and explore genomic relationships. The microarray revealed considerable genetic diversity and potential epidemiological linkages among isolates. A novel hexadecimal-based nomenclature system was introduced for standardized and scalable strain classification. Comparative analysis demonstrated that the array profiles provided greater discriminatory power and practical resolution than MLST. Receiver operating characteristic (ROC) curve analysis of 187 target genes in 220 isolates gave diagnostic sensitivity and specificity of 100%. This integrated approach offers a cost-effective, rapid, and adaptable global VRE surveillance and infection control tool. It provides a practical alternative to conventional typing systems and facilitates early detection of outbreaks and emerging clones.","PeriodicalId":8945,"journal":{"name":"BioTechniques","volume":"78 1-12","pages":"1-15"},"PeriodicalIF":2.5,"publicationDate":"2026-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"146103718","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Stabilization of human RNA and DNA in stool samples at room temperature. 室温下粪便样本中人类RNA和DNA的稳定。

IF 2.5 4区工程技术

BioTechniques Pub Date : 2025-11-01 Epub Date: 2025-08-18 DOI: 10.1080/07366205.2025.2546762

Regina Preywisch, Roger Løvlie, Moritz Eidens

{"title":"Stabilization of human RNA and DNA in stool samples at room temperature.","authors":"Regina Preywisch, Roger Løvlie, Moritz Eidens","doi":"10.1080/07366205.2025.2546762","DOIUrl":"10.1080/07366205.2025.2546762","url":null,"abstract":"This study examines the stability of human mRNA and DNA in stool samples for noninvasive gastrointestinal disease detection. While stool samples are valuable for diagnosing conditions like inflammatory disorders and colorectal cancer, mRNA instability poses significant challenges, risking false-negative results. To investigate this, 97 stool samples were treated with a specialized stabilizing solution and stored at room temperature, with analyses conducted on Day 1 and Day 15. The research aimed to improve storage protocols for enhanced reliability in mRNA diagnostics, aiding in personalized medicine and biomarker discovery. Results showed variability in total nucleic acid yields, increasing from Day 1 (mean 112 ng/µL) to Day 15 (mean 165 ng/µL), highlighting the benefits of improved homogenization and bacterial lysis. Human DNA remained stable over the 14-day period. For RNA stability, three mRNA markers were analyzed: Carcinoembryonic Antigen (CEACAM5), Prostaglandin-Endoperoxide Synthase 2 (PTGS2) and cortactin (CTTN). Both CEACAM5 (p=0.064) and PTGS2 (p=0.79) maintained stability, while CTTN showed a statistically significant but only modest reduction in expression (p < 0.0001). Overall, the stabilization buffer proved effectiveness in preserving nucleic acids and provided insights into mRNA marker stability over time.","PeriodicalId":8945,"journal":{"name":"BioTechniques","volume":" ","pages":"403-411"},"PeriodicalIF":2.5,"publicationDate":"2025-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144871263","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Artificial pre-harvest sprouting chamber with moderate humidity better simulates field sprouting for soft winter wheat. 适宜湿度的人工采前发芽室能较好地模拟软冬小麦田间发芽。

IF 2.5 4区工程技术

BioTechniques Pub Date : 2025-11-01 Epub Date: 2025-12-13 DOI: 10.1080/07366205.2025.2601538

Bryan W Penning

{"title":"Artificial pre-harvest sprouting chamber with moderate humidity better simulates field sprouting for soft winter wheat.","authors":"Bryan W Penning","doi":"10.1080/07366205.2025.2601538","DOIUrl":"10.1080/07366205.2025.2601538","url":null,"abstract":"Pre-harvest sprouting, germination of the seed on the spike, causes reduced grain quality and marketability in US wheat. Methods devised to induce and study pre-harvest sprouting vary significantly from one another in procedure, induction time, and/or measurement, and often fail to accurately reflect natural sprouting, which varies over years and locations. An artificial sprouting chamber and protocol with a shorter exposure time and relative humidity more relevant to field conditions was significantly correlated (p < 0.01) with natural pre-harvest sprouting over three years for ten wheat varieties measured by alpha amylase activity. The alpha amylase activities of ten wheat varieties tested under the developed artificial sprouting chamber and protocol were significantly correlated (p = 0.001) with those subjected to overhead irrigated field sprouting tests over three years. Four pairs of the ten varieties were genetically related but showed significantly different sprouting and alpha amylase activity. These varietal pairs may prove useful in studying the genetics of pre-harvest sprouting. While the study was performed in wheat, other susceptible crops such as rice, barley, rye, and sorghum could be tested using this method.","PeriodicalId":8945,"journal":{"name":"BioTechniques","volume":" ","pages":"367-375"},"PeriodicalIF":2.5,"publicationDate":"2025-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145740859","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Optimization of proximity labeling in endothelial cells: overcoming endogenous biotin interference and cost barriers. 内皮细胞邻近标记的优化：克服内源性生物素干扰和成本障碍。

IF 2.5 4区工程技术

BioTechniques Pub Date : 2025-11-01 Epub Date: 2026-01-29 DOI: 10.1080/07366205.2026.2619158

Ying Jiang, Kuizhi Qu, Mengjun Dai, Yan-Ning Rui, Zhen Xu

{"title":"Optimization of proximity labeling in endothelial cells: overcoming endogenous biotin interference and cost barriers.","authors":"Ying Jiang, Kuizhi Qu, Mengjun Dai, Yan-Ning Rui, Zhen Xu","doi":"10.1080/07366205.2026.2619158","DOIUrl":"https://doi.org/10.1080/07366205.2026.2619158","url":null,"abstract":"Proximity labeling has become a powerful technique for mapping protein-protein interactions under physiologically relevant conditions, with TurboID offering high enzymatic activity and rapid labeling. However, its application in endothelial systems has been limited, partly due to the presence of endogenous biotin in specialized media, which reduces labeling specificity. Here, we optimized TurboID-mediated proximity labeling in brain microvascular endothelial cells by addressing two key challenges: endogenous biotin interference and cost-effective depletion. We discovered that endothelial cell medium contains substantial biotin levels, which saturate TurboID labeling and obscure the effects of exogenous biotin. Using High Capacity NeutrAvidin™ agarose, we developed a simple and economical method to deplete endogenous biotin, reducing background biotinylation dramatically. We then defined the optimal condition for efficient labeling with minimal toxicity. Using TKS4, a scaffold protein critical for podosome formation, we validated this workflow in brain microvascular endothelial cells and confirmed the efficiency of streptavidin-based enrichment of biotinylated proteins. This study provides a validated and accessible TurboID workflow for endothelial cells, enabling more precise and cost-effective discovery of dynamic protein interaction networks relevant to vascular integrity and disease.","PeriodicalId":8945,"journal":{"name":"BioTechniques","volume":"77 11-12","pages":"1-12"},"PeriodicalIF":2.5,"publicationDate":"2025-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147472587","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0