Transgenic Research最新文献

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 T₁ 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 T₁ 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.

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.

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.

In recent years, many strategies for sustainable food systems have been launched at the national and global levels, which require better tools to monitor their progress. Subsequently, discussions on their measurements have drawn enormous attention, and various indicators have been developed. As indicators at the national level reflect policy priorities in the respective countries, it is difficult to develop adequate global indicators that accommodate different national priorities. Additionally, if we pursue only the existing dataset, we may lose the thrust of the initial objectives. However, the collection of new data can place an enormous burden on stakeholders, both developing and developed countries. These difficulties were revealed in recent negotiations for the Kunming-Montreal Global Biodiversity Framework at the Convention on Biological Diversity at the 15th Conference of the Parties. Therefore, we must reach a compromise between what we want to achieve and the resources we can share.

Given the complexity of agricultural problems, it is essential to develop acceptable solutions for various stakeholders with diverse knowledge, viewpoints, and preferences. However, European public opinion has become highly polarized, making constructive discussions on these issues difficult. We present the results of the narrative analysis of media debate on new genomic techniques. The study identified two primary narrative groups: 'precaution-focused' and 'innovation-focused.' The former emphasizes caution, potential risks, and the need for stringent regulation, while the latter highlights benefits, progress, and the promise of genome editing for sustainable agricultural practices. Within each group of narratives, several distinct narratives were identified. The research has revealed that despite the high polarization, the narratives shared important values and beliefs. Going beyond the dividing narratives and concentrating on common values can depolarize the debate and set the stage for new narratives, enabling constructive debate, concentrating on solving problems, and maximizing collective outcomes.

Whether organisms developed with the use of genome editing techniques, or food derived from such organisms, are, or should be, regulated as genetically modified organisms (GMOs) or genetically modified (GM) food, respectively, remains a subject of debate globally. Much of the discussion has been scientific and focussed on the similar genetic outcomes of some genome editing techniques and 'conventional' or natural mutagenesis. Many jurisdictions, including Australia, have considered, or are considering, how their regulatory frameworks will deal with such organisms and products. In Australia, organisms developed with site directed nuclease 1 (SDN-1, with no added template to guide homology-directed repair) are not regulated as GMOs, pursuant to exclusions in the Gene Technology Regulations 2001. The exclusion of SDN-1 organisms from regulation in Australia is sometimes misrepresented, including in scientific peer reviewed publications, as extending to all genome edited organisms. This highlights the importance for researchers, developers and other stakeholders to understand that whether genome edited organisms are, or are not, subject to regulation as GMOs in a particular jurisdiction may quintessentially be a legal question, not a scientific one.

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.

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.