Your browser does not allow saving data (e.g. private mode): bookmarks will be lost when the tab closes.
Search PubMed
Keywords, author, journal, years — results from PubMed, abstracts with Russian translation, bookmarks in your browser.
Found: 26 075
Page 53 of 1000-
Nuclease-Deficient Clustered Regularly Interspaced Short Palindromic Repeat-Based Approaches for In Vitro and In Vivo Gene Activation.
Clustered regularly interspaced short palindromic repeat (CRISPR)-based technology has been adapted to achieve a wide range of genome modifications, including transcription regulation. The focus of this review is on the application of CRISPR-based platforms such as nuclease-deficient Cas9 and Cas12a, to achieve targeted gene activation. We review studies to date that have used CRISPR-based activation technology for the elucidation of biological mechanism and disease correction, as well as its application in genetic screens as a powerful tool for high-throughput genotype-phenotype mapping. In addition to our synthesis and critical analysis of published studies, we explore key considerations for the potential clinical translation of CRISPR-based activation technology.
-
CRISPR-Cpf1 system and its applications in animal genome editing.
The clustered regularly interspaced short palindromic repeats (CRISPR) and their associated protein (Cas) system is a gene editing technology guided by RNA endonuclease. The CRISPR-Cas12a (also known as CRISPR-Cpf1) system is extensively utilized in genome editing research due to its accuracy and high efficiency. In this paper, we primarily focus on the application of CRISPR-Cpf1 technology in the construction of disease models and gene therapy. Firstly, the structure and mechanism of the CRISPR-Cas system are introduced. Secondly, the similarities and differences between CRISPR-Cpf1 and CRISPR-Cas9 technologies are compared. Thirdly, the main focus is on the application of the CRISPR-Cpf1 system in cell and animal genome editing. Finally, the challenges faced by CRISPR-Cpf1 technology and corresponding strategies are analyzed. Although CRISPR-Cpf1 technology has certain off-target effects, it can effectively and accurately edit cell and animal genomes, and has significant advantages in the preclinical research.
-
Advances in Accurate Microbial Genome-Editing CRISPR Technologies.
Previous studies have modified microbial genomes by introducing gene cassettes containing selectable markers and homologous DNA fragments. However, this requires several steps including homologous recombination and excision of unnecessary DNA regions, such as selectable markers from the modified genome. Further, genomic manipulation often leaves scars and traces that interfere with downstream iterative genome engineering. A decade ago, the CRISPR/Cas system (also known as the bacterial adaptive immune system) revolutionized genome editing technology. Among the various CRISPR nucleases of numerous bacteria and archaea, the Cas9 and Cas12a (Cpf1) systems have been largely adopted for genome editing in all living organisms due to their simplicity, as they consist of a single polypeptide nuclease with a target-recognizing RNA. However, accurate and fine-tuned genome editing remains challenging due to mismatch tolerance and protospacer adjacent motif (PAM)-dependent target recognition. Therefore, this review describes how to overcome the aforementioned hurdles, which especially affect genome editing in higher organisms. Additionally, the biological significance of CRISPR-mediated microbial genome editing is discussed, and future research and development directions are also proposed.
-
[Recent developments in enhancing the efficiency of CRISPR/Cas9- mediated knock-in in animals].
Gene-editing technology can artificially modify genetic material of targeted loci by precise insertion, deletion, or replacement in the genomic DNA. In recent years, with the developments of zinc-finger endonuclease (ZFN), transcription activator-like effector nuclease (TALEN), clustered regularly interspaced short palindromic repeats/CRISPR- associated protein 9 (CRISPR/Cas9) technologies, such precise modifications of the animal genomes have become possible. Although gene-editing tools, such as CRISPR/Cas9, can efficiently generate double-strand breaks (DSBs) in mammalian cells, the homology-directed repair (HDR) mediated knock-in (KI) efficiency is extremely low. In this review, we briefly describe the current development of gene-editing tools and summarize the recent strategies to enhance the CRISPR/Cas9- mediated KI efficiency, which will provide a reference for the generation of human disease models, research on gene therapy and livestock genetic improvement.
-
Genome Editing in Apicomplexan Parasites: Current Status, Challenges, and Future Possibilities.
Clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR-associated protein (Cas) technology has revolutionized genome editing across various biological systems, including the Apicomplexa phylum. This review describes the status, challenges, and applications of CRISPR-Cas9 editing technology in apicomplexan parasites, such as Plasmodium, Toxoplasma, Theileria, Babesia, and Cryptosporidium. The discussion encompasses successfully implemented CRISPR-Cas9-based techniques in these parasites, highlighting the achieved milestones, from precise gene modifications to genome-wide screening. In addition, the review addresses the challenges hampering efficient genome editing, including the parasites' complex life cycles, multiple intracellular stages, and the lack of robust genetic tools. It further explores the ethical and policy considerations surrounding genome editing and the future perspectives of CRISPR-Cas applications in apicomplexan parasites.
-
Therapeutic gene editing: delivery and regulatory perspectives.
Gene-editing technology is an emerging therapeutic modality for manipulating the eukaryotic genome by using target-sequence-specific engineered nucleases. Because of the exceptional advantages that gene-editing technology offers in facilitating the accurate correction of sequences in a genome, gene editing-based therapy is being aggressively developed as a next-generation therapeutic approach to treat a wide range of diseases. However, strategies for precise engineering and delivery of gene-editing nucleases, including zinc finger nucleases, transcription activator-like effector nuclease, and CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats-associated nuclease Cas9), present major obstacles to the development of gene-editing therapies, as with other gene-targeting therapeutics. Currently, viral and non-viral vectors are being studied for the delivery of these nucleases into cells in the form of DNA, mRNA, or proteins. Clinical trials are already ongoing, and in vivo studies are actively investigating the applicability of CRISPR/Cas9 techniques. However, the concept of correcting the genome poses major concerns from a regulatory perspective, especially in terms of safety. This review addresses current research trends and delivery strategies for gene editing-based therapeutics in non-clinical and clinical settings and considers the associated regulatory issues.
-
Genome-wide target specificity of CRISPR RNA-guided adenine base editors.
Adenine base editors1 enable efficient targeted adenine-to-guanine single nucleotide conversions to induce or correct point mutations in human cells, animals, and plants1-4. Here we present a modified version of Digenome-seq, an in vitro method for identifying CRISPR (clustered regularly interspaced short palindromic repeats)-induced double-strand breaks using whole-genome sequencing5-8, to assess genome-wide target specificity of adenine base editors. To produce double-strand breaks at sites containing inosines, the products of adenine deamination, we treat human genomic DNA with an adenine base editor 7.10 protein-guide RNA complex and either endonuclease V or a combination of human alkyladenine DNA glycosylase and endonuclease VIII in vitro. Digenome-seq detects adenine base editor off-target sites with a substitution frequency of 0.1% or more. We show that adenine base editor 7.10, the cytosine base editor BE3, and unmodified CRISPR-associated protein 9 (Cas9) often recognize different off-target sites, highlighting the need for independent assessments of their genome-wide specificities6. Using targeted sequencing, we also show that use of preassembled adenine base editor ribonucleoproteins, modified guide RNAs5,8-11, and Sniper/Cas9 (ref. 12) reduces adenine base editor off-target activity in human cells.
-
A CRISPR cut for messenger RNAs.
-
Chemical Modifications of CRISPR RNAs to Improve Gene-Editing Activity and Specificity.
CRISPR (clustered, regularly interspaced, short palindromic repeats) has become a cutting-edge research method and holds great potential to revolutionize biotechnology and medicine. However, like other nucleic acid technologies, CRISPR will greatly benefit from chemical innovation to improve activity and specificity for critical in vivo applications. Chemists have started optimizing various components of the CRISPR system; the present Perspective focuses on chemical modifications of CRISPR RNAs (crRNAs). As with other nucleic acid-based technologies, early efforts focused on well-established sugar and backbone modifications (2'-deoxy, 2'-F, 2'-OMe, and phosphorothioates). Some more significant alterations of crRNAs have been done using bicyclic (locked) riboses and phosphate backbone replacements (phosphonoacetates and amides); however, the range of chemical innovation applied to crRNAs remains limited to modifications that have been successful in RNA interference and antisense technologies. The encouraging results given by these tried-and-true modifications suggest that, going forward, chemists should take a bolder approach─research must aim to investigate what chemistry will have the most impact on maturing CRISPR as therapeutic and other in vivo technologies. With an eye to the future, this Perspective argues that the complexity of CRISPR presents rich unprecedented opportunities for chemists to synergize advances in synthetic methodology and structural biochemistry to rationally optimize crRNA-protein interactions.
-
CRISPR/Cas9 Immune System as a Tool for Genome Engineering.
CRISPR/Cas (clustered regularly interspaced short palindromic repeats/CRISPR-associated) adaptive immune systems constitute a bacterial defence against invading nucleic acids derived from bacteriophages or plasmids. This prokaryotic system was adapted in molecular biology and became one of the most powerful and versatile platforms for genome engineering. CRISPR/Cas9 is a simple and rapid tool which enables the efficient modification of endogenous genes in various species and cell types. Moreover, a modified version of the CRISPR/Cas9 system with transcriptional repressors or activators allows robust transcription repression or activation of target genes. The simplicity of CRISPR/Cas9 has resulted in the widespread use of this technology in many fields, including basic research, biotechnology and biomedicine.
PubMed returns only the first 10,000 results — refine the query to see the rest.