Dusan Hrckulak, Jakub Onhajzer, Michaela Krausova, Monika Stastna, Vitezslav Kriz, Lucie Janeckova, Vladimir Korinek
{"title":"Development of a new flippase-dependent mouse model for red fluorescence-based isolation of KRAS<sup>G12D</sup> oncogene-expressing tumor cells.","authors":"Dusan Hrckulak, Jakub Onhajzer, Michaela Krausova, Monika Stastna, Vitezslav Kriz, Lucie Janeckova, Vladimir Korinek","doi":"10.1007/s11248-024-00429-2","DOIUrl":"10.1007/s11248-024-00429-2","url":null,"abstract":"<p><p>Proto-oncogene KRAS, GTPase (KRAS) is one of the most intensively studied oncogenes in cancer research. Although several mouse models allow for regulated expression of mutant KRAS, selective isolation and analysis of transforming or tumor cells that produce the KRAS oncogene remains a challenge. In our study, we present a knock-in model of oncogenic variant KRAS<sup>G12D</sup> that enables the \"activation\" of KRAS<sup>G12D</sup> expression together with production of red fluorescent protein tdTomato. Both proteins are expressed from the endogenous Kras locus after recombination of a transcriptional stop box in the genomic DNA by the enzyme flippase (Flp). We have demonstrated the functionality of the allele termed RedRas (abbreviated Kras<sup>RR</sup>) under in vitro conditions with mouse embryonic fibroblasts and organoids and in vivo in the lung and colon epithelium. After recombination with adenoviral vectors carrying the Flp gene, the Kras<sup>RR</sup> allele itself triggers formation of lung adenomas. In the colon epithelium, it causes the progression of adenomas that are triggered by the loss of tumor suppressor adenomatous polyposis coli (APC). Importantly, cells in which recombination has successfully occurred can be visualized and isolated using the fluorescence emitted by tdTomato. Furthermore, we show that KRAS<sup>G12D</sup> production enables intestinal organoid growth independent of epidermal growth factor (EGF) signaling and that the KRAS<sup>G12D</sup> function is effectively suppressed by specific inhibitor MRTX1133.</p>","PeriodicalId":23258,"journal":{"name":"Transgenic Research","volume":"34 1","pages":"9"},"PeriodicalIF":2.7,"publicationDate":"2025-01-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11717838/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142955700","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":"Exploring genetic mapping and co-expression patterns to illuminate significance of Tbx20 in cardiac biology.","authors":"Dezhong Zhang, Xiao Shang, Quanquan Ji, Li Niu","doi":"10.1007/s11248-024-00423-8","DOIUrl":"https://doi.org/10.1007/s11248-024-00423-8","url":null,"abstract":"<p><p>The transcription factor Tbx20 is integral to heart development and plays a significant role in various cardiac diseases. Despite its established importance, the regulatory mechanisms and functional significance of Tbx20 remain incompletely understood. To elucidate these mechanisms, we initially conducted eQTL mapping to identify genetic loci associated with Tbx20 expression in heart tissue from BXD mice. Co-expression and enrichment analyses revealed pathways linked to Tbx20, including dilated cardiomyopathy, hypertrophic cardiomyopathy, and FoxO signaling. Additionally, protein-protein interaction studies identified essential cardiac proteins, such as Myl2 and Myl7, along with upstream regulators like Mef2c. To validate our bioinformatic findings, we performed quantitative reverse transcription polymerase chain reaction (qRT-PCR) to assess the relative mRNA expression levels of TBX20 and Mef2c in the heart tissues of BXD mice compared to their parental strains (B6 and D2). Our results demonstrated significant up-regulation of both TBX20 and Mef2c in the BXD group relative to the parental strains. Conversely, both genes were down-regulated in B6, D2, Control, and Treatment groups when compared to BXD mice. These findings confirm the predicted regulatory roles of TBX20 and Mef2c in cardiac development as suggested by our initial analyses.This study not only reinforces the critical role of Tbx20 in cardiac gene regulation but also highlights its potential as a therapeutic target for cardiovascular disorders. Further investigations into Tbx20 and its interactions will enhance our understanding of heart biology and contribute to the development of targeted therapies for heart diseases.</p>","PeriodicalId":23258,"journal":{"name":"Transgenic Research","volume":"34 1","pages":"5"},"PeriodicalIF":2.7,"publicationDate":"2025-01-07","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142955702","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}
Kei Takamoto, Kaori Inazu, Shuichi Nakai, Koichi Inoue, Mai Tsuda
{"title":"Do confined field trials add value for the environment risk assessment of genetically modified Brassica napus L. in Japan?","authors":"Kei Takamoto, Kaori Inazu, Shuichi Nakai, Koichi Inoue, Mai Tsuda","doi":"10.1007/s11248-024-00425-6","DOIUrl":"10.1007/s11248-024-00425-6","url":null,"abstract":"<p><p>The environmental risk assessment (ERA) of genetically modified (GM) crops in Japan requires collecting data from a comparative study of a GM and non-GM control in an in-country confined field trial (CFT). This in-country CFT requirement is used to address concerns that differences in the local environmental conditions may lead to differences in growth and/or risks of GM crops. However, this requirement for in-country CFT has recently been exempted for certain GM maize and GM cotton traits, and instead CFT data from other countries are used to inform the ERA of these GM events. However, in-country CFTs continue to be required for GM B. napus. Our objective is to assess whether using B. napus as a host crop increases the potential for differences between GM B. napus and conventional B. napus that may have an impact on biodiversity occurring only under the Japanese environment. In this paper agronomic data was compiled from seven local CFTs of GM B. napus events to assess the potential for differences between GM and non-GM B. napus for three key areas; competitiveness, potential to produce harmful substances, and outcrossing. Considering these elements, the need for conducting CFTs locally for ERA of future GM B. napus traits is discussed. The assessment concluded that conducting CFT locally is not necessary for GM B. napus events if traits do not bring competitive advantage or produce harmful substances only under Japanese environment.</p>","PeriodicalId":23258,"journal":{"name":"Transgenic Research","volume":"34 1","pages":"6"},"PeriodicalIF":2.7,"publicationDate":"2025-01-07","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11706835/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142955701","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}
Rita Verma, Anshu Sahu, Rajan Kumar Gupta, Indraneel Sanyal
{"title":"Sonication-assisted Rhizobium radiobacter-mediated genetic transformation of Indian Lotus (Nelumbo nucifera Gaertn.).","authors":"Rita Verma, Anshu Sahu, Rajan Kumar Gupta, Indraneel Sanyal","doi":"10.1007/s11248-024-00427-4","DOIUrl":"https://doi.org/10.1007/s11248-024-00427-4","url":null,"abstract":"<p><p>This study aimed to develop a reliable and efficient genetic transformation method for the ornamental Indian Lotus (Nelumbo nucifera Gaertn.) using the sonication-assisted Rhizobium radiobacter-mediated transformation technique. To conduct the transformation, shoot apical meristem explants were infected with Rhizobium radiobacter (synonym Agrobacterium tumefaciens) strain LBA 4404 containing a binary vector pBI121 that harbours the GUS reporter gene (uidA) and kanamycin resistance gene nptII for plant selection. To improve the transformation efficiency, we optimized parameters such as bacterial cell density, sonication duration, infection time, co-cultivation duration, acetosyringone concentration, cefotaxime, and kanamycin concentrations. Sonication treatment at 42 kHz for 90 s recorded the highest transformation efficiency. The selection of regenerated plantlets was performed on a kanamycin-supplemented selection medium. The putative transformants showed GUS expression in the leaves and petioles. The presence of the GUS gene was also confirmed in the putative transformants through PCR, with the appearance of the expected amplicon size of 520 bp. The presence of nptII was confirmed by PCR in the putatively transformed plants with an amplicon size of 530 bp. The maximum regeneration frequency obtained was 72.66%, and the highest transformation efficiency achieved was 9.0% in the Indian Lotus.</p>","PeriodicalId":23258,"journal":{"name":"Transgenic Research","volume":"34 1","pages":"4"},"PeriodicalIF":2.7,"publicationDate":"2025-01-07","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142955663","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}
Chunying Gong, Junwen Ai, Yong Liu, Xingjian He, Hong Xue, Chaohua Jia, Zhuohua Chen, Hanfu Xu, Rongpeng Liu, Yong Yang
{"title":"Transgenic overexpression of UDP glycosyltransferase gene UGT41A3 induces resistance to nucleopolyhedrovirus in Bombyx mori.","authors":"Chunying Gong, Junwen Ai, Yong Liu, Xingjian He, Hong Xue, Chaohua Jia, Zhuohua Chen, Hanfu Xu, Rongpeng Liu, Yong Yang","doi":"10.1007/s11248-024-00422-9","DOIUrl":"https://doi.org/10.1007/s11248-024-00422-9","url":null,"abstract":"<p><p>Bombyx mori nuclear polyhedrosis, caused by B. mori nucleopolyhedrovirus (BmNPV), threatens sericulture seriously. To explore strategies for controlling it, the UDP glycosyltransferase gene UGT41A3 (BmUGT41A3) was targeted. UGT is involved in exogenous substances detoxification and endogenous biomass regulation in insects. Early embryos of the BmNPV-sensitive variety 'HYB' were used to obtain the transgenic line HYB-UGT41A3, overexpressing BmUGT41A3 under the IE1 promoter. qPCR results revealed that, compared with the wild-type control 'HYB', BmUGT41A3 was upregulated during the individual developmental stages of HYB-UGT41A3 from silkworm eggs to fifth-instar larvae; peak expression was observed in the third-instar larvae, which presented the most significantly upregulated expression. Individual-tissues qPCR results revealed that BmUGT41A3 expression was highest in the hemocytes of HYB-UGT41A3, followed by the midgut, whereas expression in HYB was very low. Gradient feeding of BmNPV on HYB-UGT41A3 and control 'HYB' larvae on the first day of the second-instar stage. The results revealed that the LC<sub>50</sub> of HYB-UGT41A3 reached 4.040 × 10<sup>7</sup> particles/mL, which was 20-fold greater than that of HYB. The decrease in the BmNPV load was more significant in HYB-UGT41A3 than in HYB at 48 h after viral inoculation. These results indicate BmUGT41A3 overexpression inhibits BmNPV proliferation and improve resistance to BmNPV in B. mori.</p>","PeriodicalId":23258,"journal":{"name":"Transgenic Research","volume":"34 1","pages":"2"},"PeriodicalIF":2.7,"publicationDate":"2024-12-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142910893","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}
Xue-Ni Zhong, Jun-Jie Peng, Meng-Yao Wang, Xiu-Li Yang, Li Sun
{"title":"Overexpression of NAC transcription factors from Eremopyrum triticeum promoted abiotic stress tolerance.","authors":"Xue-Ni Zhong, Jun-Jie Peng, Meng-Yao Wang, Xiu-Li Yang, Li Sun","doi":"10.1007/s11248-024-00428-3","DOIUrl":"https://doi.org/10.1007/s11248-024-00428-3","url":null,"abstract":"<p><p>Eremopyrum triticeum is a typical spring ephemeral species, which in China mainly distributed in the desert regions of northern Xinjiang, and play an important role in the desert ecosystems. E. triticeum has several adaptive characteristics such as short growth rhythms, high photosynthetic efficiency, high seed production, drought and salt resistance. However, the molecular regulatory mechanism of E. triticeum in responses to abiotic stress resistance is still unknown. In this study, two NAC-like transcription factor-encoding genes, EtNAC1 and EtNAC2, were isolated from E. triticeum. The predicted EtNAC1 and EtNAC2 proteins possess a typical NAC DNA-binding domain at the N-terminal region. The qRT-PCR analysis showed that EtNAC1 and EtNAC2 were highly expressed in mature roots of E. triticeum, and were significantly up-regulated under drought, high salt and abscisic acid (ABA) stresses. Subcellular localization analysis in onion epidermal cells revealed that EtNAC1 and EtNAC2 were located in the nucleus. Expression of EtNAC1 and EtNAC2 in yeast cells improved the survival rate of yeast under low temperature, H<sub>2</sub>O<sub>2</sub>, high drought and salt stresses. Overexpression of EtNAC1 and EtNAC2 in Arabidopsis thaliana conferred enhanced tolerance to drought and salt stresses, increased ABA sensitivity, and transgenic plants showed higher proline (Pro) content, but lower malondialdehyde content, lower chlorophyll leaching, lower water loss rate and stomatal aperture (width/length) than WT plants. In conclusion, EtNAC1 and EtNAC2 play important roles in abiotic stress responses of E. triticeum, which might have significant potential in crop molecular breeding for abiotic stress tolerance.</p>","PeriodicalId":23258,"journal":{"name":"Transgenic Research","volume":"34 1","pages":"3"},"PeriodicalIF":2.7,"publicationDate":"2024-12-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142910892","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}
Peng Xu, Shaojuan Lai, Bo Yin, Chenyu Yao, Xiaolin Gu, Jilei Huang, Yufei Hu
{"title":"Interplay between Agrobacterium T-DNA and backbone DNA in transgenic plant cells.","authors":"Peng Xu, Shaojuan Lai, Bo Yin, Chenyu Yao, Xiaolin Gu, Jilei Huang, Yufei Hu","doi":"10.1007/s11248-024-00424-7","DOIUrl":"https://doi.org/10.1007/s11248-024-00424-7","url":null,"abstract":"<p><p>Agrobacterium-mediated transformation of plants often results in the integration of multiple copies of T-DNA and backbone DNA from binary vectors into the host genome. However, the interplay between T-DNA and backbone DNA remains elusive. In this study, 70.8% of T<sub>1</sub> Arabidopsis transformants exhibited integration of both T-DNA and backbone DNA, and no cases of only backbone integration were observed. To elucidate the integration patterns, we employed bulk-genome resequencing in Arabidopsis and identified 20 integration sites across 10 T<sub>1</sub> transgenic plants, most of which were flanked by left borders of T-DNA at both ends. On average, each integration site contained 6.3 copies of T-DNA and 2.65 copies of backbone DNA. The junction structures between T-DNA and the backbone were highly variable, revealing a previously underappreciated frequency of readthrough at both the left and right borders. Transient expression studies in Nicotiana benthamiana leaves demonstrated that T-DNA and backbone DNA were simultaneously transferred into transformed cells, although the backbone DNA had lower copy numbers than T-DNA. These findings suggest a close relationship between T-DNA and backbone DNA during their transfer and integration, thus offering new insights into the mechanism underlying Agrobacterium-mediated transformation.</p>","PeriodicalId":23258,"journal":{"name":"Transgenic Research","volume":"34 1","pages":"1"},"PeriodicalIF":2.7,"publicationDate":"2024-12-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142910891","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}
Transgenic ResearchPub Date : 2024-12-01Epub Date: 2024-10-21DOI: 10.1007/s11248-024-00414-9
Urs Niggli
{"title":"Sustainability requires the integration of farmer knowledge, scientific advancements, and comprehensive innovation.","authors":"Urs Niggli","doi":"10.1007/s11248-024-00414-9","DOIUrl":"10.1007/s11248-024-00414-9","url":null,"abstract":"<p><p>This viewpoint paper emphasises the need to diversify food production methods to simultaneously combat hunger and reduce environmental problems. The recommendations of the UN Food System Summit 2021 relate primarily to (i) the conservation of natural ecosystems, (ii) the sustainable management of existing agricultural land while increasing productivity and (iii) the restoration of already degraded land. Europe in particular faces unique challenges, such as reducing pollution and promoting organic farming up to 25 percent of the agricultural land area while maintaining food production. Ongoing efforts aim to create a transparent, fair and multi-level regulatory framework to support the Green Deal. The implementation of the Corporate Sustainability Reporting Directive (CSRD), which will sooner or later affect a larger proportion of European farmers, should support the transition. Science and innovation play a central role in this, as they are the cornerstones on which sustainable food systems are built. It is imperative that farmers actively participate in the co-design processes and utilise their wealth of experience and creativity to drive these innovations forward. A crucial aspect of the transition to sustainability is changing consumption patterns to limit food waste and reduce meat consumption. While this transition is essential, it is not without its formidable challenges. Diversification of agriculture, encompassing a spectrum of established techniques, is touted as a promising approach to achieving sustainability without sacrificing productivity. Furthermore, integrating truly sustainable agricultural practices with cutting-edge innovations, including new genomic techniques, has the potential to be a transformative solution.</p>","PeriodicalId":23258,"journal":{"name":"Transgenic Research","volume":" ","pages":"563-569"},"PeriodicalIF":2.7,"publicationDate":"2024-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142475604","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}
Transgenic ResearchPub Date : 2024-12-01Epub Date: 2024-08-06DOI: 10.1007/s11248-024-00397-7
Oli Watson, Sadiye Hayta
{"title":"Precision breeding in agriculture and food systems in the United Kingdom.","authors":"Oli Watson, Sadiye Hayta","doi":"10.1007/s11248-024-00397-7","DOIUrl":"10.1007/s11248-024-00397-7","url":null,"abstract":"<p><p>In recent years there have been major advances in precision breeding technologies, such as gene editing, that offer promising solutions to revolutionise global crop production and tackle the pressing issues in food systems. The UK has leading expertise in genomics, and research is already taking place to develop crops with improved resilience to climate change, resistance to disease and less reliance on chemical inputs. In March 2023, the Genetic Technology (Precision Breeding) Act received Royal Assent and passed into UK law. It provides a framework from which to build more proportionate regulations for plants and animals made using genetic technologies which contain genetic changes that could also arise through traditional breeding-known as 'Precision Bred Organisms'. New legislation and the utilization of UK world-leading research could help to enhance the efficiency of breeding systems and enable the development of plants and animals that are healthier, better for the environment and more resilient to climate change.</p>","PeriodicalId":23258,"journal":{"name":"Transgenic Research","volume":" ","pages":"539-544"},"PeriodicalIF":2.7,"publicationDate":"2024-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11655596/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141894338","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}