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CRISPR-Cas Gene Editing Technology in Potato.
Potato (Solanum tuberosum L.) is one of the most important food crops in the world, ranking fourth after rice, maize, and wheat. Potatoes are exposed to biotic and abiotic environmental factors, which lead to economic losses and increase the possibility of food security threats in many countries. Traditional potato breeding faces several challenges, primarily due to its genetic complexity and the time-consuming nature of the process. Therefore, gene editing-CRISPR-Cas technology-allows for more precise and rapid changes to the potato genome, which can speed up the breeding process and lead to more effective varieties. In this review, we consider CRISPR-Cas technology as a potential tool for plant breeding strategies to ensure global food security. This review summarizes in detail current and potential technological breakthroughs that open new opportunities for the use of CRISPR-Cas technology for potato breeding, as well as for increasing resistance to abiotic and biotic stresses, and improving potato tuber quality. In addition, the review discusses the challenges and future perspectives of the CRISPR-Cas system in the prospects of the development of potato production and the regulation of gene-edited crops in different countries around the world.
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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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How to respond to CRISPR babies.
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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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[The CRISPR/Cas system: a genome editing tool to develop animal models of viral infections].
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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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Gasdermin E as a potential target and biomarker for CRISPR-Cas9-based cancer therapy.
Gasdermin E (GSDME), a protein pivotal in mediating pyroptosis, has gained significant attention due to its role in cancer pathogenesis and its potential as a therapeutic target. The advent of CRISPR-Cas9, a precise genome editing tool, has revolutionized cancer therapy by enabling the manipulation of GSDME expression and function. This review explores the interplay of GSDME and CRISPR-Cas9 in cancer, emphasizing GSDME's unique mechanism of cleavage-dependent pore formation in the cell membrane and its emerging applications as both a therapeutic target and a diagnostic biomarker. We discuss the potential and challenges of using GSDME-induced pyroptosis as a therapeutic strategy and how can enhance its efficacy and specificity. We conclude by highlighting promising future research directions in this emerging field.
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Single-molecule perspectives of CRISPR/Cas systems: target search, recognition, and cleavage.
CRISPR/Cas systems have emerged as powerful tools for gene editing, nucleic acid detection, and therapeutic applications. Recent advances in single-molecule techniques have provided new insights into the DNA-targeting mechanisms of CRISPR/ Cas systems, in particular, Types I, II, and V. Here, we review how single-molecule approaches have expanded our understanding of key processes, namely target search, recognition, and cleavage. Furthermore, we focus on the dynamic behavior of Cas proteins, including PAM site recognition and R-loop formation, which are crucial to ensure specificity and efficiency in gene editing. Additionally, we discuss the conformational changes and interactions that drive precise DNA cleavage by different Cas proteins. This mini review provides a comprehensive overview of CRISPR/Cas molecular dynamics, offering conclusive insights into their broader potential for genome editing and biotechnological applications. [BMB Reports 2025; 58(1): 8-16].
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Synthetically modified guide RNA and donor DNA are a versatile platform for CRISPR-Cas9 engineering.
Chemical modification of the gRNA and donor DNA has great potential for improving the gene editing efficiency of Cas9 and Cpf1, but has not been investigated extensively. In this report, we demonstrate that the gRNAs of Cas9 and Cpf1, and donor DNA can be chemically modified at their terminal positions without losing activity. Moreover, we show that 5' fluorescently labeled donor DNA can be used as a marker to enrich HDR edited cells by a factor of two through cell sorting. In addition, we demonstrate that the gRNA and donor DNA can be directly conjugated together into one molecule, and show that this gRNA-donor DNA conjugate is three times better at transfecting cells and inducing HDR, with cationic polymers, than unconjugated gRNA and donor DNA. The tolerance of the gRNA and donor DNA to chemical modifications has the potential to enable new strategies for genome engineering.
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