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Future Perspectives of Prime Editing for the Treatment of Inherited Retinal Diseases.
Inherited retinal diseases (IRD) are a clinically and genetically heterogenous group of diseases and a leading cause of blindness in the working-age population. Even though gene augmentation therapies have shown promising results, they are only feasible to treat a small number of autosomal recessive IRDs, because the size of the gene is limited by the vector used. DNA editing however could potentially correct errors regardless of the overall size of the gene and might also be used to correct dominant mutations. Prime editing is a novel CRISPR/Cas9 based gene editing tool that enables precise correction of point mutations, insertions, and deletions without causing double strand DNA breaks. Due to its versatility and precision this technology may be a potential treatment option for virtually all genetic causes of IRD. Since its initial description, the prime editing technology has been further improved, resulting in higher efficacy and a larger target scope. Additionally, progress has been achieved concerning the size-related delivery issue of the prime editor components. This review aims to give an overview of these recent advancements and discusses prime editing as a potential treatment for IRDs.
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Harnessing CRISPR-Cas technology for insect pest control: current advances and future perspectives.
Insect pests pose a significant threat to agriculture, public health, and global biodiversity, prompting the development of innovative control strategies. CRISPR-Cas technology, a revolutionary genome-editing tool, has emerged as a promising approach to tackle these challenges. This review comprehensively explores the current advances in using CRISPR-Cas systems for insect pest control, highlighting key methodologies, successes, and potential hurdles. The fundamental principles of CRISPR-Cas technology, emphasizing its precision and versatility in gene editing, have been outlined. We then delve into various pest-control applications, such as gene drive systems designed to spread genetic modifications through pest populations, thereby reducing their reproductive capacity or increasing their susceptibility to environmental factors. Successful applications of CRISPR-based strategies in insect species such as Aedes aegypti, where gene editing has been used to suppress vector populations and reduce disease transmission potential, and Spodoptera frugiperda, where targeted gene disruption has improved pest management studies, are also highlighted. Furthermore, the ecological and ethical implications of deploying CRISPR-Cas based interventions are discussed, including concerns about unintended consequences, resistance development, and impacts on non-target species. Regulatory frameworks and public acceptance are also examined as critical components for the successful implementation of these technologies. The review concludes with a forward-looking perspective on the future of CRISPR-Cas in pest control, considering advancements in delivery methods, precision editing, and integration with other pest management strategies. We propose potential research directions and highlight the need for interdisciplinary collaboration to optimize the use of CRISPR-Cas technology for sustainable, effective insect pest management. This article serves as a comprehensive resource for researchers, policymakers, and stakeholders, providing a detailed overview of the current state of CRISPR-Cas technology in insect pest control and its future potential to transform pest management practices.
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How to respond to CRISPR babies.
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From the first human gene-editing to the birth of three-parent baby.
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LION: a simple and rapid method to achieve CRISPR gene editing.
The RNA-guided CRISPR-Cas9 technology has paved the way for rapid and cost-effective gene editing. However, there is still a great need for effective methods for rapid generation and validation of CRISPR/Cas9 gRNAs. Previously, we have demonstrated that highly efficient generation of multiplexed CRISPR guide RNA (gRNA) expression array can be achieved with Golden Gate Assembly (GGA). Here, we present an optimized and rapid method for generation and validation in less than 1 day of CRISPR gene targeting vectors. The method (LION) is based on ligation of double-stranded gRNA oligos into CRISPR vectors with GGA followed by nucleic acid purification. Using a dual-fluorescent reporter vector (C-Check), T7E1 assay, TIDE assay and a traffic light reporter assay, we proved that the LION-based generation of CRISPR vectors are functionally active, and equivalent to CRISPR plasmids generated by traditional methods. We also tested the activity of LION CRISPR vectors in different human cell types. The LION method presented here advances the rapid functional validation and application of CRISPR system for gene editing and simplified the CRISPR gene-editing procedures.
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CRISPR-Mediated Gene Editing for Inducing Thermosensitive Genic Male Sterility and Sheath Blight Resistance in Rice.
Recent advances in genome editing enable the researchers to focus more and more on the ability to manipulate genomes at specific sites. Efficient methods for genome editing further promote gene discovery and functional gene analyses in model plants as well as the introduction of novel desired agricultural traits in important species. CRISPR/Cas9 technology enables precise genetic modification through the creation of double-strand breaks in a target region and the generation of desired alterations during the repair process. In this chapter, we describe the cloning strategy, transformation protocols, triparental mating procedure, and characterization of genome-edited genetic male sterile mutants and sheath blight disease-resistant mutant plants.
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Efficient genome editing in cultured cells and embryos of Debao pig and swamp buffalo using the CRISPR/Cas9 system.
Myostatin (MSTN), a protein encoded by growth differentiation factor 8 (GDF8), is primarily expressed in skeletal muscle and negatively regulates the development and regeneration of muscle. Accordingly, myostatin-deficient animals exhibit a double-muscling phenotype. The CRISPR/Cas9 system has proven to be an efficient genome-editing tool and has been applied to gene modification in cells from many model organisms such as Drosophila melanogaster, zebrafish, mouse, rat, sheep, and human. Here, we edited the GDF8 gene in fibroblasts and embryos of Debao pig and swamp buffalo using the CRISPR/Cas9 system. The CRISPR/Cas9-mediated mutation efficiency in fibroblasts was as high as 87.5% in pig and 78.9% in buffalo. We then obtained single-cell clones with mutations at the specific sites of the GDF8 gene by screening with G418 in fibroblasts of pig and buffalo. In addition, the frequencies of Cas9/gRNA-mediated mutations were at 36 and 25% in the intracytoplasmic sperm injection embryos of pig and in vitro fertilization embryos of buffalo, respectively. Our work demonstrates that the Cas9/gRNA system is a highly efficient and fast tool for genome editing in cultured cells and embryos of Debao pig and swamp buffalo. These results can be helpful for the establishment of a new animal strain that can generate more meat.
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Synergizing gene editing and cellular agriculture for a sustainable and healthy food future.
The intersection of gene editing and cellular agriculture is transforming food production by offering sustainable, ethical alternatives to conventional agriculture. Cellular agriculture uses tissue engineering and fermentation technologies to produce animal-free food, whereas gene editing tools like CRISPR-Cas9 optimize cellular efficiency, nutritional value, and sustainability. While some researchers emphasize the environmental and food security benefits, others raise concerns about high costs, regulatory challenges, and consumer acceptance. This paper critically examines existing literature, compares breakthroughs and controversies, and provides an expert perspective on the challenges and opportunities in gene-edited cellular agriculture. By tackling key scientific, economic, regulatory, and ethical issues, this article presents a roadmap for responsibly advancing these technologies and integrating them into global food systems. To our knowledge, this is the first work to explore how gene editing and cellular agriculture can be synergized to advance sustainability, food security, and global health.
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Impact of Gene Editing Tools, Like CRISPR/Cas9, on the Public Health Response to Disease Outbreaks.
The purpose of this communication is to explore the implications of genome editing techniques, such as CRISPR/Cas9, on public health-related responses to outbreaks of disease. The recent commercialization of genome editing techniques makes the creation and release of genetically altered pathogens a much easier task, increasing the possibility to the point of needing discussion. Three areas need to be addressed: predictions concerning potential genetic alterations, predictions and implications concerning the release of genetically altered pathogens, and the short- and long-term implications of the release of genetically altered pathogens. Full discourse on these topics among professionals in the area of public health will help to combat harm from the use of any genetically altered biologic weapons. The topics covered here include a review of the CRISPR/Cas9 gene editing technique, including a discussion of which possibilities utilize genome editing. We then address predictions about the application of gene alterations in the context of bioweapons. We discuss a few basic concepts about the evolution of an intentionally released genetically altered organism based on circumstances and patterns gleaned from observing nature in the hope that this will aid in the public health response to bioterrorism attack. (Disaster Med Public Health Preparedness. 2017;11:155-159).
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[Are the Argonauts lost at sea?].
A novel gene editing procedure based on a nuclease from the Argonaute protein family was described in mid-2016 and appeared to provide significant advantages over the now widely used CRISPR-Cas9 system. Attempts by numerous groups to use this technique have however been unsuccessful; several negative reports have been published in addition to many accounts of failure found in the "grey literature". It is unclear at this point whether this reflects an (unknown) critical experimental factor or hints at data misinterpretation, possibly even at outright fabrication of results.
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