BMC Biotechnology最新文献

{"title":"An engineered hypoxia-response promoter for human umbilical cord-derived mesenchymal stem cell-based therapeutics.","authors":"Song Yang, Weizhong Zhuang, Lishi Zhou, Weiwei Kong, Wanwan Zou, Qikun Zhu, Enze Bian, Bin Lin, Jianzheng Cen, Qiang Gao, Jimei Chen","doi":"10.1186/s12896-025-00993-3","DOIUrl":"10.1186/s12896-025-00993-3","url":null,"abstract":"<p><p>Myocardial infarction, characterized by insufficient blood supply to the heart, leads to ischemia and hypoxia of myocardial tissues, causing injury and decreased cardiac function. Despite improvements in pharmaceutical and interventional therapies, it remains a leading cause of death worldwide. Human umbilical cord mesenchymal stem cells (hUC-MSCs) play an important role in the repair of infarcted myocardium by promoting angiogenesis, reducing inflammation, secreting growth factors and cytokines. However, the harsh hypoxic microenvironment of infarcted myocardial tissue poses a threat to the survival and function of transplanted hUC-MSCs. In this study, we modified the candidate gene promoter of hUC-MSCs under hypoxic conditions and created a promoter that can respond quickly under hypoxic conditions. We found that the modified promoter significantly promoted the transcription efficiency as hypoxia time increased. This indicates that the engineered hypoxia-response promoter can effectively drive gene expression in a hypoxic environment. Furthermore, the transcription efficiency of the modified promoter under normoxic conditions is lower than that of common promoters in eukaryotic organisms, suggesting that this effect can improve the efficacy and safety of hUC-MSC-based myocardial infarction treatment by ensuring that cells function effectively in the damaged hypoxic area.</p>","PeriodicalId":8905,"journal":{"name":"BMC Biotechnology","volume":"25 1","pages":"59"},"PeriodicalIF":3.5,"publicationDate":"2025-07-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12220557/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144537901","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":"Unconventional semi-solid cultivation enhances cytochalasins production by the Colombian fungus Xylaria sp. CM-UDEA-H199.","authors":"Daniela Valencia-Revelo, Esteban Charria-Girón, Katharina Schmidt, Silke Reinecke, Aida M Vasco-Palacios, Theresia Stradal, Yasmina Marin-Felix, Nelson H Caicedo-Ortega, Sherif S Ebada","doi":"10.1186/s12896-025-00978-2","DOIUrl":"10.1186/s12896-025-00978-2","url":null,"abstract":"<p><p>Fungal species of the order Xylariales, particularly those from tropical and untapped areas like the Amazon region, denote an intriguing reservoir of biodiversity and chemically varied metabolites. Based on this potential and by implementing the One Strain Many Compounds (OSMAC) approach, herein we have cultivated a Colombian Xylaria strain in several liquid, solid or semi-solid media, under different nutrient compositions and culture conditions. Metabolomic studies of Xylaria sp. CM-UDEA-H199 across these conditions led to the isolation of diverse metabolites. Six compounds were purified from rice (BRFT) cultures, identified as griseofulvin (1), xylaropyrones B/C (2/3), akolitserin (4), hypoxylin A (5), and (-)-(R)-5-(methoxycarbonyl)mellein (6). Three compounds were isolated from liquid YM cultivation: 2-hexylidene-3-methylsuccinic acid (7), its 4-methyl ester (8), and akoenic acid (9). Notably, cultivation in the newly designed semi-solid (S-BRFT) medium significantly altered the metabolome, leading to the predominant production of cytochalasins, with five derivatives (10-14) purified and structurally characterized.Among the isolated cytochalasins, compound 12 was identified as a previously undescribed natural diepoxy derivative of cytochalasin D. Structure elucidation of all isolated compounds was achieved based on their MS and comprehensive 1D/2D NMR analyses in addition to comparisons with the reported literature. Compounds 4-6, 10 and 11 revealed mild antifungal activity, while compounds (1, 5, 6, 8, 10, 11, 13 and 14) exhibited cytotoxic activity, with hypoxylin A (5) being the most potent, displaying IC₅₀ values in the nanomolar range. In cellulo studies revealed that the epimerization at C-5 of cytochalasin D (10) backbone, as in hypoxylin A (5), neither affected its activity nor reversibility on actin dynamics. However, the epoxylated variant of 10, cytochalasin R (14), enhanced actin activity accompanied by reduced cytotoxicity compared to 5 and 10. The occurrence of diverse epoxy-substituted cytochalasins suggests that specific biosynthetic enzymes were activated in response to the applied fermentation conditions. These findings provide a basis for further bioprocess optimization strategies aimed at enhancing cytochalasan production, a chemical class recognized for its promising bioactivities in recent decades.</p>","PeriodicalId":8905,"journal":{"name":"BMC Biotechnology","volume":"25 1","pages":"57"},"PeriodicalIF":3.5,"publicationDate":"2025-07-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12220629/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144537993","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":"A simplified two-plasmid system for orthogonal control of mammalian gene expression using light-activated CRISPR effector.","authors":"Shruthi S Garimella, Shiaki A Minami, Anusha N Khanchandani, Justin C Abad Santos, Susannah R Schaffer, Priya S Shah","doi":"10.1186/s12896-025-00994-2","DOIUrl":"10.1186/s12896-025-00994-2","url":null,"abstract":"<p><strong>Background: </strong>Optogenetic systems use light-responsive proteins to control gene expression, ion channels, protein localization, and signaling with the \"flip of a switch\". One such tool is the light activated CRISPR effector (LACE) system. Its ability to regulate gene expression in a tunable, reversible, and spatially resolved manner makes it attractive for many applications. However, LACE relies on delivery of four separate components on individual plasmids, which can limit its use. Here, we optimize LACE to reduce the number of plasmids needed to deliver all four components.</p><p><strong>Results: </strong>The two-plasmid LACE (2pLACE) system combines the four components of the original LACE system into two plasmids. Following construction, the behavior of 2pLACE was rigorously tested using optogenetic control of enhanced green fluorescent protein (eGFP) expression as a reporter. Using human HEK293T cells, we optimized the ratio of the two plasmids, measured activation as a function of light intensity, and determined the frequency of the light to activate the maximum fluorescence. Overall, the 2pLACE system showed a similar dynamic range, tunability, and activation kinetics as the original four plasmid LACE (4pLACE) system. Interestingly, 2pLACE also had less variability in activation signal compared to 4pLACE. We also demonstrate the optimal LACE system also depends on cell type. In mouse myoblast C2C12 cells, 2pLACE displayed less variability compared to 4pLACE, similar to HEK293T cells. However, 2pLACE also had a smaller dynamic range in C2C12 cells compared to 4pLACE.</p><p><strong>Conclusions: </strong>This simplified system for optogenetics will be more amenable to biotechnology applications where variability needs to be minimized. By optimizing the LACE system to use fewer plasmids, 2pLACE becomes a flexible tool in multiple research applications. However, the optimal system may depend on cell type and application.</p>","PeriodicalId":8905,"journal":{"name":"BMC Biotechnology","volume":"25 1","pages":"58"},"PeriodicalIF":3.5,"publicationDate":"2025-07-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12220775/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144537900","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":"Assessing bone regeneration potential of 3D scaffold-free cell pellets from periodontal ligament and bone marrow stem cells.","authors":"Xiang Liang, Yuxin Gong, Le Bai, Sina Ahmadi, Ming Yu, Zhou Zhang, Siya Fang, Fangfang Xu, Weiqi Wang, Junbo Tu, Sijia Na","doi":"10.1186/s12896-025-00983-5","DOIUrl":"10.1186/s12896-025-00983-5","url":null,"abstract":"<p><strong>Background: </strong>Bone defects pose a significant clinical challenge in clinical treatment, where stem cell-based tissue engineering strategies have emerged as a promising approach for bone regeneration. Notably, accumulating evidence suggests that scaffold-free three-dimensional cell pellets demonstrate therapeutic potential through direct implantation into bone defects area.</p><p><strong>Methods: </strong>Two types of stem cells were isolated from bone and tooth, then cultured separately. Surface markers (CD34, CD45, CD90, CD105, CD146, STRO-1) were analyzed by flow cytometry. Bone marrow-derived and dental-derived CPs were cultured in ascorbic acid-supplemented growth medium. Histological morphology from CS to CP was examined through H&E staining. Cell pellets' biological properties were assessed in vitro via immunofluorescence, qPCR, and Western blot, and bone regeneration was evaluated using rat calvarial defect models.</p><p><strong>Results: </strong>We demonstrated comparable morphology and immunophenotype between BMSCs and PDLSCs. H&E staining and TUNEL assays revealed tightly organized histological structures and low apoptosis rates in 5-day cultured cell pellets. Immunofluorescence analysis showed no significant differences in COL-1 or early osteogenic marker ALP between PDLSC-CP and BMSC-CP; however, BMSC-CP exhibited higher osteoblast-related BSP expression, whereas PDLSC-CP displayed elevated bone remodeling-associated OPN levels. Consistent trends were observed in qPCR and Western blot analyses. In rat calvarial defect models, both CP types induced significant bone formation, with BMSC-CP demonstrating enhanced osteogenic capacity compared to PDLSC-CP.</p><p><strong>Conclusion: </strong>Both BMSC-CPs and PDLSC-CPs demonstrate osteogenic potential in scaffold-free 3D environments, whereas standardized controls and mechanistic investigations are required to establish their distinct therapeutic advantages.</p>","PeriodicalId":8905,"journal":{"name":"BMC Biotechnology","volume":"25 1","pages":"55"},"PeriodicalIF":3.5,"publicationDate":"2025-07-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12210542/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144537903","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":"Central position of histidine in the sequence of designed alternating polarity peptides enhances pH-responsive assembly with DNA.","authors":"Razieh Taghizadeh Pirposhteh, Nasir Mohajel, Arash Arashkia, Kayhan Azadmanesh, Mohammadali Masoumi","doi":"10.1186/s12896-025-00976-4","DOIUrl":"10.1186/s12896-025-00976-4","url":null,"abstract":"<p><strong>Background: </strong>Self-assembling peptides hold great promise for medical applications, particularly as carriers for gene delivery, but their potential remains unrealized due to a limited understanding of how amino acid sequence positioning affects their properties. In this study, we designed and evaluated two alternating polarity peptides, RFH (RFRHRHRFR) and RHF (RHRFRFRHR), differing only in the position of their histidine and phenylalanine residues, to investigate the impact of sequence variation on pH-responsive DNA co-assembly and transfection efficiency.</p><p><strong>Results: </strong>Both peptides formed stable co-assemblies with DNA at neutral pH. However, RHF retained its co-assembling ability at acidic pH, as confirmed by gel retardation studies. Coarse-grained molecular dynamics simulations further supported these findings, showing a reduced affinity of RFH for DNA and a sharper decrease in DNA binding when its histidine residues were protonated. Morphological analysis revealed that both co-assemblies underwent structural transitions with increasing N/P ratios, though their sizes and morphologies differed significantly. Biological studies demonstrated that both peptides achieved a higher transfection efficiency in 293T and HeLa cells compared to R₉, with a lower cytotoxicity than polyethyleneimine. Notably, RFH exhibited superior transfection performance at lower N/P ratios compared to RHF, likely due to its distinct histidine and phenylalanine arrangement and its pH-responsive co-assembly behavior with DNA.</p><p><strong>Conclusions: </strong>These findings highlight the importance of histidine positioning within peptide sequences for tuning pH-responsiveness and optimizing DNA co-assembly and transfection efficiency. The results provide valuable insights for the rational design of efficient, safe, and pH-responsive peptide-based gene delivery systems.</p>","PeriodicalId":8905,"journal":{"name":"BMC Biotechnology","volume":"25 1","pages":"54"},"PeriodicalIF":3.5,"publicationDate":"2025-07-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12210498/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144537985","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":"High yield purification of an isoleucine zipper-modified CD95 ligand for efficient cell apoptosis initiation and with biotin or DNA-oligomer binding domain to probe ligand functionalization effects.","authors":"Xiaoyue Shang, Nina Bartels, Johann Moritz Weck, Sabine Suppmann, Jérôme Basquin, Gajen Thaventhiran, Amelie Heuer-Jungemann, Cornelia Monzel","doi":"10.1186/s12896-025-00986-2","DOIUrl":"10.1186/s12896-025-00986-2","url":null,"abstract":"<p><strong>Background: </strong>Cluster of differentiation 95 (CD95/Fas/Apo1) as part of the Tumor-necrosis factor (TNF) receptor family is a prototypic trigger of the 'extrinsic' apoptotic pathway and its activation by the trimeric ligand CD95L is of high interest. However, CD95L, when presented in solution, exhibits a low efficiency to induce apoptosis signaling in human cells.</p><p><strong>Results: </strong>Here, we design a recombinant CD95L exhibiting an isoleucine zipper (IZ) motif at the N-terminus for stabilization of the trimerized CD95L and demonstrate its high apoptosis initiation efficiency. This efficiency is further enhanced by antibody-mediated crosslinking of IZ-CD95L.A cysteine amino acid fused behind the IZ is used as a versatile coupling site for bionanotechnological applications or for the development of biomedical assays. A fast, cheap, and efficient production of CD95L via the HEK293T secretory expression system is presented, along with CD95L affinity purification and functionalization. We verified the biological activity of the purified protein and identified a stabilized trimeric CD95L structure as the most potent inducer of apoptosis signaling.</p><p><strong>Conclusions: </strong>The workflow and the findings reported here will streamline a wide array of future low- or high-throughput TNF-ligand screens, and their modification towards improving apoptosis induction efficiency and, potentially, anticancer therapy.</p>","PeriodicalId":8905,"journal":{"name":"BMC Biotechnology","volume":"25 1","pages":"64"},"PeriodicalIF":3.5,"publicationDate":"2025-07-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12219679/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144537988","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":"Self-assembling T7 phage syringes with modular genomes for targeted delivery of penicillin against β-lactam-resistant Escherichia coli.","authors":"Hyunjin Shim","doi":"10.1186/s12896-025-01003-2","DOIUrl":"10.1186/s12896-025-01003-2","url":null,"abstract":"<p><p>Bacteriophages are promising alternative antimicrobial agents due to their high specificity for host bacteria and minimal immunogenicity in humans. However, their therapeutic application is limited by their nature as biological entities, with potential evolutionary consequences. In this study, we address these challenges by repurposing only the structural components of bacteriophages as vesicles to deliver antibiotics directly into the cytoplasm of bacterial hosts. This approach is based on two key hypotheses: first, antibiotics such as β-lactams remain effective against resistant bacteria if injected directly into the cytoplasm, bypassing resistance mechanisms; second, phage structures can be synthesized and self-assembled in vitro using modular genomes and cell-free protein expression to carry small molecules as cargo. To test these hypotheses, we utilized T7 phages and penicillin-resistant Escherichia coli as a model system. First, we designed the T7 phage genome into a modular format containing only the genes encoding structural components and synthesized the gene fragments via de novo gene synthesis. These phage structures were then rebooted in vitro using cell-free protein expression in the presence of penicillin G, allowing the antibiotics to be incorporated as cargo during the self-assembly process. Finally, we tested the antimicrobial activity of these antibiotic-loaded phage syringes against penicillin-resistant E. coli. The results demonstrate that phage syringes effectively reduce the host population compared to negative controls, including free penicillin and water. This study highlights the potential of using phage structures as antibiotic delivery vehicles, offering a novel strategy to overcome both the limitations of small-molecule antibiotics and phage therapy.</p>","PeriodicalId":8905,"journal":{"name":"BMC Biotechnology","volume":"25 1","pages":"63"},"PeriodicalIF":3.5,"publicationDate":"2025-07-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12220360/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144537992","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":"Purification, characterization, and anti-cancer activity of methionine gamma-lyase from a native strain of Pseudomonas mosselii for human cancer treatment.","authors":"Matin Nasirian, Mohsen Mobini-Dehkordi, Pegah Khosravian","doi":"10.1186/s12896-025-00995-1","DOIUrl":"10.1186/s12896-025-00995-1","url":null,"abstract":"<p><strong>Aims: </strong>Methionine gamma-lyase (MGL) specifically targets L-methionine-dependent cancer cells, making it a promising candidate for anti-cancer drug development. This study aims to purify and characterize L-methioninase from Pseudomonas mosselii and evaluate its potential anti-cancer properties.</p><p><strong>Methods and results: </strong>MGL was purified through heat treatment, ion exchange chromatography, and gel filtration, achieving a 6-fold purification and a 58.43% recovery rate. The enzyme displayed an activity of 61.16 U/mg and had a molecular weight of 48 kDa. Optimal activity was observed at a pH of 6 and temperatures ranging from 30 to 37℃. Kinetic studies revealed a Km value of 8.458 mM and a Vmax of 0.2702 U/mL/min for L-methionine. The anti-cancer effects of MGL were tested on MCF-7, MOLT-4, HepG-2, and U87MG cell lines. MTT assays demonstrated significant anti-cancer activity, inducing apoptosis in cancer cells while sparing normal fibroblasts. Real-time PCR results demonstrated decreased expression of BCL-2 and increased expression of caspase-3. This further confirms that apoptosis is enhanced by the use of gamma-lyase enzyme and methionine restriction in cancer cells.</p><p><strong>Conclusions: </strong>MGL shows promise as a targeted treatment for L-methionine-dependent cancers by selectively inducing apoptosis in cancer cells. Its specific action and effective purification establish MGL as a potential therapeutic candidate.</p>","PeriodicalId":8905,"journal":{"name":"BMC Biotechnology","volume":"25 1","pages":"61"},"PeriodicalIF":3.5,"publicationDate":"2025-07-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12220622/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144537991","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":"Lactococcus lactis I7 isolated from traditional Italian cheese making: a biotechnological integrated platform.","authors":"Alberto Alfano, Darshankumar Parecha, Alessandra Fusco, Vittoria Savio, Ida De Chiara, Lidia Muscariello, Giovanna Donnarumma, Chiara Schiraldi","doi":"10.1186/s12896-025-00997-z","DOIUrl":"10.1186/s12896-025-00997-z","url":null,"abstract":"<p><strong>Background: </strong>There is growing interest in newly isolated lactic acid bacteria from traditional sources. In this study, a Lactococcus lactis strain identified and isolated from natural whey starter cultures of cow milk for the production of artisanal cheeses was cultivated in optimized vegan grade medium to assess its growing ability and metabolic fingerprint. In fact, traditionally fermented dairy products are considered nutritionally complete and possibly functional to human health with a number of benefits not only to the gastro-intestinal tract but also in a systemic manner.</p><p><strong>Results: </strong>In fed batch experiments, we achieved 14 g/L dry cell weight and 1.9∙10¹⁰ viable colony forming unit increasing the values of experiment threefold and twofold respect to the batch processes, respectively. Additionally, the lactic acid (LA) production was quantified, and a maximal concentration of 78 g/L was achieved, which is approximately five-fold corresponding to the batch experiment results. This is in agreement with kinetic modeling of LA inhibition studies that highlighted a halved growth rate (µ) at 35 ± 5 g/L of LA whilst the growth blockage occurred at about 80 g/L. The samples obtaining after ultrafiltration processes and tested on three different pathogens, Enterococcus faecalis, Salmonella enterica subsp. enterica serovar Typhimurium and enteroinvasive Escherichia coli, gave a reduction on viability by 6-9 log on average.</p><p><strong>Conclusion: </strong>This paper focused its attention on optimization of fermentation conditions (in particular a vegan grade media using Design of Experiment approach) using fed-batch and microfiltration processes increasing the production of biomass and bioactive molecules, with respect to the batch processes. At the end of the fed-batch fermentation, a downstream process based on membranes was performed in order to obtain bioactive molecules that proved antimicrobial activity against intestinal/food spoilage pathogens.</p>","PeriodicalId":8905,"journal":{"name":"BMC Biotechnology","volume":"25 1","pages":"62"},"PeriodicalIF":3.5,"publicationDate":"2025-07-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12219981/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144537989","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":"Mother transformer: A High-Throughput, Cost-Effective in Planta Hairy Root Transformation Method for Cannabis.","authors":"Ladan Ajdanian, Davoud Torkamaneh","doi":"10.1186/s12896-025-00990-6","DOIUrl":"10.1186/s12896-025-00990-6","url":null,"abstract":"<p><strong>Background: </strong>Hairy root (HR) transformation assays mediated by Agrobacterium rhizogenes, both in vitro and ex vitro, are essential tools in plant biotechnology and functional genomics. These assays can be significantly influenced by various factors, which ultimately can enhance the efficiency. In this study, we optimized a two-step ex vitro HR transformation method using the actual mother plant combined with the RUBY system and compared with existing methods.</p><p><strong>Results: </strong>The two-step ex vitro method proved more efficient than both the one-step ex vitro and in vitro methods, with the highest transformation efficiency of 90% observed in the actual plant. This technique also demonstrated a faster and less complicated approach, reducing time to achieve massive transgenic HR formation by 9-29 days compared to other methods.</p><p><strong>Conclusions: </strong>A novel, quicker, less complicated, and more efficient two-step transformation method for cannabis has been established, presenting a significantly lower risk of contamination. This protocol is particularly interesting to produce secondary metabolites using the CRISPR/Cas system in cannabis. We anticipate that this method will facilitate substantial time savings by rapidly producing hundreds of transformed samples.</p>","PeriodicalId":8905,"journal":{"name":"BMC Biotechnology","volume":"25 1","pages":"60"},"PeriodicalIF":3.5,"publicationDate":"2025-07-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12219872/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144537990","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}