Transgenic Research最新文献

{"title":"Correction to: WIF1 causes dysfunction of heart in transgenic mice.","authors":"Dan Lu, Wei Dong, Xu Zhang, Xiongzhi Quan, Dan Bao, Yingdong Lu, Lianfeng Zhang","doi":"10.1007/s11248-024-00403-y","DOIUrl":"10.1007/s11248-024-00403-y","url":null,"abstract":"","PeriodicalId":23258,"journal":{"name":"Transgenic Research","volume":" ","pages":"535-536"},"PeriodicalIF":2.7,"publicationDate":"2024-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141983369","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Mutagenesis on a complex mouse genetic background by site-specific nucleases.","authors":"Benjamin Davies, Lucy Trelfa, Victoria S Rashbrook, Edward Drydale, Rachel Martin, Boyan Bai, Jedrzej Golebka, Daniel Stephen Biggs, Keith M Channon, Shoumo Bhattacharya, Gillian Douglas","doi":"10.1007/s11248-024-00399-5","DOIUrl":"10.1007/s11248-024-00399-5","url":null,"abstract":"<p><p>Mouse models with complex genetic backgrounds are increasingly used in preclinical research to accurately model human disease and to enable temporal and cell-specific evaluation of genetic manipulations. Backcrossing mice onto these complex genetic backgrounds takes time and leads to significant wastage of animals. In this study, we aimed to evaluate whether site-specific nucleases could be used to generate additional genetic mutations in a complex genetic background, using the REVERSA mouse model of atherosclerosis, a model harbouring four genetically altered alleles. The model is comprised of a functional null mutation in the Ldlr gene in combination with a ApoB100 allele, which, after high-fat diet, leads to the rapid development of atherosclerosis. The regression of the pathology is achieved by inducible knock-out of the Mttp gene. Here we report an investigation to establish if microinjection of site-specific nucleases directly into zygotes prepared from the REVERSA could be used to investigate the role of the ATP binding cassette transporter G1 (ABCG1) in atherosclerosis regression. We show that using this approach we could successfully generate two independent knockout lines on the REVERSA background, both of which exhibited the expected phenotype of a significant reduction in cholesterol efflux to HDL in bone marrow-derived macrophages. However, loss of Abcg1 did not impact atherosclerosis regression in either the aortic root or in aortic arch, demonstrating no important role for this transporter subtype. We have demonstrated that site-specific nucleases can be used to create genetic modifications directly onto complex disease backgrounds and can be used to explore gene function without the need for laborious backcrossing of independent strains, conveying a significant 3Rs advantage.</p>","PeriodicalId":23258,"journal":{"name":"Transgenic Research","volume":" ","pages":"415-426"},"PeriodicalIF":2.7,"publicationDate":"2024-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11588839/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141860987","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":"Characterizing a lethal CAG-ACE2 transgenic mouse model for SARS-CoV-2 infection using Cas9-enhanced nanopore sequencing.","authors":"Alexander Smirnov, Artem Nurislamov, Galina Koncevaya, Irina Serova, Evelyn Kabirova, Eduard Chuyko, Ekaterina Maltceva, Maxim Savoskin, Daniil Zadorozhny, Victor A Svyatchenko, Elena V Protopopova, Oleg S Taranov, Stanislav S Legostaev, Valery B Loktev, Oleg Serov, Nariman Battulin","doi":"10.1007/s11248-024-00413-w","DOIUrl":"10.1007/s11248-024-00413-w","url":null,"abstract":"<p><p>The SARS-CoV-2 pandemic has underscored the necessity for functional transgenic animal models for testing. Mouse lines with overexpression of the human receptor ACE2 serve as the common animal model to study COVID-19 infection. Overexpression of ACE2 under a strong ubiquitous promoter facilitates convenient and sensitive testing of COVID-19 pathology. We performed pronuclear microinjections using a 5 kb CAG-ACE2 linear transgene construct and identified three founder lines with 140, 72, and 73 copies, respectively. Two of these lines were further analyzed for ACE2 expression profiles and sensitivity to SARS-CoV-2 infection. Both lines expressed ACE2 in all organs analyzed. Embryonic fibroblast cell lines derived from transgenic embryos demonstrated severe cytopathic effects following infection, even at low doses of SARS-CoV-2 (0,1-1.0 TCID₅₀). Infected mice from the two lines began to show COVID-19 clinical signs three days post-infection and succumbed between days 4 and 7. Histological examination of lung tissues from terminally ill mice revealed severe pathological alterations. To further characterize the integration site in one of the lines, we applied nanopore sequencing combined with Cas9 enrichment to examine the internal transgene concatemer structure. Oxford Nanopore sequencing (ONT) is becoming the gold standard for transgene insert characterization, but it is relatively inefficient without targeted region enrichment. We digested genomic DNA with Cas9 and gRNA against the ACE2 transgene to create ends suitable for ONT adapter ligation. ONT data analysis revealed that most of the transgene copies were arranged in a head-to-tail configuration, with palindromic junctions being rare. We also detected occasional plasmid backbone fragments within the concatemer, likely co-purified during transgene gel extraction, which is a common occurrence in pronuclear microinjections.</p>","PeriodicalId":23258,"journal":{"name":"Transgenic Research","volume":" ","pages":"453-466"},"PeriodicalIF":2.7,"publicationDate":"2024-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142354510","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Promoter of COR2-like gene is a stress inducible regulatory region in banana.","authors":"Sanjana Negi, Nikita Mahashabde, Subham Bhakta, Sudhir Singh, Himanshu Tak","doi":"10.1007/s11248-024-00405-w","DOIUrl":"10.1007/s11248-024-00405-w","url":null,"abstract":"<p><p>A promoter is a crucial component in driving the expression of a transgene of interest for biotechnological applications in crop improvement and thus characterization of varied regulatory regions is essential. Here, we identified the promoter of COR2-like (codeinone reductase-like) from banana and characterized its tissue specific and stress inducible nature. MusaCOR2-like of banana is closely related to COR2 and CHR (chalcone reductase) sequences from different plant species and contains signature sequences including a catalytic tetrad typical of proteins with aldo-keto reductase activity. Transcript level of MusaCOR2-like was strongly induced in response to drought, salinity and exposure of signaling molecules such as abscisic acid, methyl-jasmonate and salicylic acid. Induction of MusaCOR2-like under stress strongly correlated with the presence of multiple cis-elements associated with stress responses in the P_{MusaCOR2-like} sequence isolated from Musa cultivar Rasthali. Transgenic tobacco lines harbouring P_{MusaCOR2-like}-GUS displayed visible GUS expression in vascular tissue of leaves and stem while its expression was undetectable in roots under control conditions. Exposure to drought, salinity and cold strongly induced GUS expression from P_{MusaCOR2-like}-GUS in transgenic tobacco shoots in a window period of 3H to 12H. Applications of salicylic acid, methyl-jasmonate, abscisic acid and ethephon also activate GUS in transgenic shoots at different period, with salicylic acid and abscisic acid being the stronger stimulants of P_{MusaCOR2-like}. Using P_{MusaCOR2-like}-GUS fusion and expression profiling, the current study sheds insights into a complex regulation of COR2-like, one of the least studied genes of secondary metabolite pathway in plants.</p>","PeriodicalId":23258,"journal":{"name":"Transgenic Research","volume":" ","pages":"399-413"},"PeriodicalIF":2.7,"publicationDate":"2024-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11588891/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142112329","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":"CRISPR/Cas12a ribonucleoprotein mediated editing of tryptophan 2,3-dioxygenase of Spodoptera frugiperda.","authors":"Dhawane Yogi, Karuppannasamy Ashok, Cholenahalli Narayanappa Anu, Thalooru Shashikala, Chalapathy Pradeep, Chikmagalur Nagaraja Bhargava, Madhusoodanan Sujatha Parvathy, M N Jithesh, Maligeppagol Manamohan, Girish Kumar Jha, Ramasamy Asokan","doi":"10.1007/s11248-024-00406-9","DOIUrl":"10.1007/s11248-024-00406-9","url":null,"abstract":"<p><p>In insect genome editing CRISPR/Cas9 is predominantly employed, while the potential of several classes of Cas enzymes such as Cas12a largely remain untested. As opposed to Cas9 which requires a GC-rich protospacer adjacent motif (PAM), Cas12a requires a T-rich PAM and causes staggered cleavage in the target DNA, opening possibilities for multiplexing. In this regard, the utility of Cas12a has been shown in only a few insect species such as fruit flies and the silkworm, but not in non-model insects such as the fall armyworm, Spodoptera frugiperda, a globally important invasive pest that defies most of the current management methods. In this regard, a more recent genetic biocontrol method known as the precision-guided sterile insect technique (pgSIT) has shown successful implementation in Drosophila melanogaster, with certain thematic adaptations required for application in agricultural pests. However, before the development of a controllable gene drive for a non-model species, it is important to validate the activity of Cas12a in that species. In the current study we have, for the first time, demonstrated the potential of Cas12a by editing an eye color gene, tryptophan 2,3-dioxygenase (TO) of S. frugiperda by microinjecting ribonucleoprotein complex into pre-blastoderm (G0) eggs. Analysis of G0 mutants revealed that all five mutants (two male and three female) exhibited distinct edits consisting of both deletion and insertion events. All five edits were further validated through in silico modeling to understand the changes at the protein level and further corroborate with the range of eye-color phenotypes observed in the present study.</p>","PeriodicalId":23258,"journal":{"name":"Transgenic Research","volume":" ","pages":"369-381"},"PeriodicalIF":2.7,"publicationDate":"2024-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142112328","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Public perception of folate-biofortified genetically modified lettuce varieties in Brazil.","authors":"Thaís de Moura Cipriano, Maria Thereza Macedo Pedroso, Isis Amanda de Paula Nunes, Lídia Nascimento Queiroz, Francisco José Lima Aragão","doi":"10.1007/s11248-024-00400-1","DOIUrl":"10.1007/s11248-024-00400-1","url":null,"abstract":"<p><p>Lettuce is one of the most widely consumed vegetables in the world, commonly eaten fresh in salads, sandwiches, wraps, and as a garnish in various dishes. Consequently, it is a very promising vehicle to deliver vitamins, such as folate (vitamin B9), to a specific population using biofortified varieties generated by conventional or molecular breeding. A new genetically modified lettuce was generated with increased folate content. However, some issues related to public perception regarding this technology should still be evaluated. The aim of this study was to analyze whether consumers are willing to accept a folate-biofortified GM lettuce that could become available to the Brazilian market. A questionnaire involving several issues regarding lettuce consumption was answered by 2,391 people from almost all Brazilian states. When informed that the folic acid biofortified lettuce is a transgenic plant, 46.1% of respondents stated that they would eat it and 30.5% stated that it would be a possibility. This study demonstrated that if there is any explanation regarding the advantage in relation to the use of biotechnology, like enrichment with folic acid, the number of people who accept it increases.</p>","PeriodicalId":23258,"journal":{"name":"Transgenic Research","volume":" ","pages":"359-368"},"PeriodicalIF":2.7,"publicationDate":"2024-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141894339","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Next-generation CRISPR technology for genome, epigenome and mitochondrial editing.","authors":"Cia-Hin Lau, Qing-Le Liang, Haibao Zhu","doi":"10.1007/s11248-024-00404-x","DOIUrl":"10.1007/s11248-024-00404-x","url":null,"abstract":"<p><p>The application of rapidly growing CRISPR toolboxes and methods has great potential to transform biomedical research. Here, we provide a snapshot of up-to-date CRISPR toolboxes, then critically discuss the promises and hurdles associated with CRISPR-based nuclear genome editing, epigenome editing, and mitochondrial editing. The technical challenges and key solutions to realize epigenome editing in vivo, in vivo base editing and prime editing, mitochondrial editing in complex tissues and animals, and CRISPR-associated transposases and integrases in targeted genomic integration of very large DNA payloads are discussed. Lastly, we discuss the latest situation of the CRISPR/Cas9 clinical trials and provide perspectives on CRISPR-based gene therapy. Apart from technical shortcomings, ethical and societal considerations for CRISPR applications in human therapeutics and research are extensively highlighted.</p>","PeriodicalId":23258,"journal":{"name":"Transgenic Research","volume":" ","pages":"323-357"},"PeriodicalIF":2.7,"publicationDate":"2024-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142000730","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"OsHRZ1 negatively regulates rice resistant to Magnaporthe oryzae infection by targeting OsVOZ2.","authors":"Jia-Ying Sun, Zeng-Ran Zhou, Yu-Qi Wang, Dong-Yu Zhu, Dian-Rong Ma","doi":"10.1007/s11248-024-00415-8","DOIUrl":"10.1007/s11248-024-00415-8","url":null,"abstract":"<p><p>Rice blast disease caused by Magnaporthe oryzae significantly reduces yield production. Blast resistance is closely associated with iron (Fe) status, but the mechanistic basis linking iron status to immune function in rice remains largely unknown. Here, iron-binding haemerythrin RING ubiquitin ligases OsHRZ1 was confirmed to play key roles in iron-mediated rice blast resistance. The expression of OsHRZ1 was suppressed by M. oryzae inoculation and high iron treatment. Both mutants of OsHRZ1 enhanced rice resistance to M. oryzae. OsPR1a was up-regulated in OsHRZ1 mutants. Yeast two-hybrid, bimolecular fluorescence complementation, and Co-IP assay results indicated that OsHRZ1 interacts with Vascular Plant One Zinc Finger 2 (OsVOZ2) in the nucleus. Additionally, the vitro ubiquitination assay indicated that OsHRZ1 can ubiquitinate OsVOZ2 and mediate the degradation of OsVOZ2. The mutants of OsVOZ2 showed reduced resistance to M. oryzae and down-regulated the expression of OsPR1a. Yeast one-hybrid, EMSA, and dual-luciferase reporter assay results indicated that OsVOZ2 directly binds to the promoter of OsPR1a, activating its expression. In summary, OsHRZ1 plays an important role in rice disease resistance by mediated degradation of OsVOZ2 thus shaping PR gene expression dynamics in rice cells. This highlights an important link between iron signaling and rice pathogen defenses.</p>","PeriodicalId":23258,"journal":{"name":"Transgenic Research","volume":" ","pages":"489-501"},"PeriodicalIF":2.7,"publicationDate":"2024-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142354511","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Long-term (10-year) monitoring of transposon-mediated transgenic cattle.","authors":"Soo-Young Yum, Bae Young Choi, Gyeong-Min Gim, Kyeong-Hyeon Eom, Seong-Beom Lee, Daehyun Kim, Euntaek Lim, Do-Yoon Kim, Seong-Eun Heo, Donghwan Shim, Goo Jang","doi":"10.1007/s11248-024-00401-0","DOIUrl":"10.1007/s11248-024-00401-0","url":null,"abstract":"<p><p>The production of transgenic animals using non-viral methods has raised questions regarding their long-term health and genomic stability. In this study, we evaluated these aspects in transgenic cattle over ten years, using transposon-mediated gene transfer. Our longitudinal analysis included a comprehensive health assessment and whole-genome DNA resequencing. We found no significant alterations in physiological parameters or health complications in transposon-mediated transgenic cattle that exceeded 10 years of age. Genomic analysis revealed that the rates of somatic mutations and copy number variations in transgenic cattle were comparable to those in non-transgenic cattle. Furthermore, structural variants were infrequent, suggesting that transposon-mediated gene insertion did not compromise genomic integrity. These findings highlight the viability of transposon systems for generating transgenic livestock, potentially expanding their applications in agriculture and biotechnology. This study contributes significantly to our understanding of the long-term implications of transgenesis in large animals and supports the safety and stability of this method.</p>","PeriodicalId":23258,"journal":{"name":"Transgenic Research","volume":" ","pages":"503-512"},"PeriodicalIF":2.7,"publicationDate":"2024-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11588892/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142081705","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":"Functional analysis of root-preferential oil palm metallothionein promoter in tobacco.","authors":"Subhi Siti Masura, Noor Azmi Shaharuddin, Mat Yunus Abdul Masani, Kuang-Lim Chan, Eng-Ti Leslie Low, Pek-Lan Chan, Abdul Rahman Siti Rahmah, Nadzirah Amiruddin, Mohd Puad Abdullah, Azzreena Mohamad Azzeme, Ghulam Kadir Ahmad Parveez, Omar Abd Rasid","doi":"10.1007/s11248-024-00396-8","DOIUrl":"10.1007/s11248-024-00396-8","url":null,"abstract":"<p><p>Root-specific or preferential promoters are essential to genetically modify plants with beneficial root traits. We have characterised the promoter from an oil palm metallothionein gene (EgMT) and performed a serial 5' deletion analysis to identify the region(s) essential for transgenes expression in roots. Stable functional characterisation of tobacco transgenic lines using the T₁ generation showed that a deletion construct, designated as RSP-2D (1107 bp), directed strong GUS expression at all stages of root development, particularly in mature roots. Other constructs, RSP-2A (2481 bp) and RSP-2C (1639 bp), drove GUS expression in roots with an intensity lower than RSP-2D. The promoter activity was also detectable in seed pods and immature seeds, albeit at lower levels than CaMV35S. The promoter activity may also be induced by wounding as intact GUS staining was observed at the flower- and leaf-cutting sites of T₁ samples carrying either RSP-2C or RSP-2D constructs. The promoter sequence contains cis-acting elements that may act as negative regulators and be responsible for root specificity. The results further indicated that the 5' UTR and ATATT sequences are essential for strong promoter activity. This study highlights the potential of RSP-2D promoter as a tool for modifying root traits through genetic engineering.</p>","PeriodicalId":23258,"journal":{"name":"Transgenic Research","volume":" ","pages":"383-397"},"PeriodicalIF":2.7,"publicationDate":"2024-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141907807","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}