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: 26 168
Page 720 of 1000-
Graphite Bio Pauses Lead Gene Editing Program in Sickle Cell Disease.
-
As CRISPR-Cas adoption soars, summit calls for genome editing oversight.
-
Deleting ku80 improves the efficiency of targeted gene editing in Neospora caninum.
CRISPR/Cas9 technology has been widely used for gene editing in organisms. Gene deletion of the ku80/ku70 complex can improve the efficiency of gene replacement in Arabidopsis thaliana, Cryptococcus neoformans, and Toxoplasma gondii, which remained elusive in Neospora caninum. Here, we knock out the ku80 gene in Nc1 strain by using CRISPR/Cas9, detect the growth rate and virulence of NcΔku80. Then we compare the efficiency of gene replacements between NcΔku80 and Nc1 strains by transfected with the same HA-tagged plasmids, and the percentage of HA-tagged parasites was investigated by IFA. The results showed that gene targeting efficiency was increased in the NcΔku80 strain via double crossover at several genetic loci, but its growth rate and virulence were unaffected. In conclusion, the NcΔku80 strain can be used as an effective strain for rapid gene editing of N. caninum.
-
Monkey survives for two years after gene-edited pig-kidney transplant.
-
In Vivo Investigation of Gene Function in Muscle Stem Cells by CRISPR/Cas9-Mediated Genome Editing.
Skeletal muscle satellite cells (SCs) are adult stem cells responsible for muscle development and injury-induced muscle regeneration. Functional elucidation of intrinsic regulatory factors governing SC activity is constrained partially by the technological limitations in editing SCs in vivo. Although the power of CRISPR/Cas9 in genome manipulation has been widely documented, its application in endogenous SCs remains largely untested. Our recent study generates a muscle-specific genome editing system leveraging the Cre-dependent Cas9 knockin mice and AAV9-mediated sgRNAs delivery, which allows gene disruption in SCs in vivo. Here, we illustrate the step-by-step procedure for achieving efficient editing using the above system.
-
The gene-therapy revolution risks stalling if we don't talk about drug pricing.
-
Generation of Cashmere Goats Carrying an EDAR Gene Mutant Using CRISPR-Cas9-Mediated Genome Editing.
In recent years, while the use of the clustered regularly interspaced short palindromic repeat (CRISPR)-CRISPR-associated protein 9 (Cas9) (CRISPR-Cas9) system for targeted genome editing has become a research hotspot, it has, to date, not proved adequate for genome editing in large mammals, such as goats. In this study, two opposite single-guide RNAs (sgRNAs) were designed for complete EDAR gene targeting in Cashmere goats, and co-transfected with a plasmid encoding Cas9 into goat fibroblasts. Among the 89 cell lines obtained through the cultivation of clonal cell lines, 62 were positive for EDAR gene targeting. Nine types of mutations were identified by sequencing analysis, and the mutation efficiency was 69.7%. Using one of these cell lines, EDAR gene-targeted Cashmere goat embryos were prepared by somatic cell cloning. Developed embryos were transferred to 79 Cashmere goat recipients, and, after a gestation period of five months six male EDAR gene-targeted Cashmere goats were born. Although only two of these goats survived, they had abnormal primary hair follicles and no hair on the top of their heads, which are the distinctive features of the EDAR gene-targeted Cashmere goats. Thus, this study provides a valuable animal model for future studies on EDAR gene-related phenotypes and hair follicle growth and development and shows that the CRISPR-Cas9 system can be used to edit genes in large mammals.
-
Study on the framework of ATP energy cycle system in Escherichia coli.
The high mortality rate associated with single-use CRISPR-Cas9 in Escherichia coli limits its application. Recently, new CRISPR-based techniques for E.coli gene editing have emerged. Research aims to develop a system for rapid, marker-free, multi-site, and multi-copy genome editing in E.coli to advance synthetic biology. ATP, essential for energy in living organisms, plays a crucial role in various metabolic processes. To reduce the cost of ATP-requiring reactions, it is crucial to identify and efficiently express genes in ATP synthesis pathway. This study identified a single ppk gene (No.8) capable of completing the cyclic reaction. Using MUCICAT technology, the ppk gene (No.8) was inserted into various positions and copy numbers in the E.coli genome, resulting in different activity levels. The findings suggest that the difficulty of inserting the ppk gene (No.8) into the genome follows this order: IS186 < 8array < IS186 + 8array < IS1. A single genome insertion can mimic plasmid expression level. This study explores promoter competition and offers solutions, inspiring researchers in constructing the AMP-ATP cycle system in E.coli. KEY POINTS: • The single ppk gene (No.8) can regenerate the AMP-ATP cycle, crucial for ATP-dependent reactions. • Inserting the ppk gene (No.8) into the cr5 site of the E.coli genome achieves expression levels comparable to the pET29a plasmid. • The expression level of the ppk gene (No.8) is not significantly affected by its copy number in the E.coli genome.
-
A Simple and Efficient Procedure for Developing Mouse Germline Stem Cell Lines with Gene Knock-in via CRISPR/Cas9 Technology.
Cultured mammalian spermatogonial stem cells (SSCs), also known as germline stem cells (GSCs), hold great promise for applications such as fertility preservation, gene therapy, and animal breeding, particularly in conjunction with accurate gene editing. Although the in vitro development of mouse GSC (mGSC) lines, and gene-targeting procedures for such lines, were initially established about two decades ago, it remains challenging for beginners to efficiently accomplish these tasks, partly because mGSCs proliferate more slowly and are more resistant to lipid-mediated gene transfection than pluripotent stem cells (PSCs). Meanwhile, methods for mGSC culture and gene editing have been evolving constantly to become simpler and more efficient. Here, we describe how to develop mGSC lines from small mouse testis samples and how to carry out gene knock-in in these cells using CRISPR/Cas9 technology, detailing three basic protocols that constitute a streamlined procedure. Using these simple and efficient procedures, site-specific knock-in mGSC lines can be obtained in 3 months. We hope that these protocols will help researchers use genetically modified GSCs to explore scientific questions of interest and to accumulate experience for application to GSC research in other mammalian species. © 2024 Wiley Periodicals LLC. Basic Protocol 1: Establishment of mouse GSCs lines from small testicular samples Basic Protocol 2: Preparation of plasmids for gene knock-in using the CRISPR/Cas9 system Basic Protocol 3: Establishment of gene knock-in mGSC lines by electroporation gene delivery.
-
CRISPR-mediated modification of DNA methylation pattern in the new era of cancer therapy.
In the last 2 decades, a wide variety of studies have been conducted on epigenetics and its role in various cancers. A major mechanism of epigenetic regulation is DNA methylation, including aberrant DNA methylation variations such as hypermethylation and hypomethylation in the promoters of critical genes, which are commonly detected in tumors and mark the early stages of cancer development. Therefore, epigenetic therapy has been of special importance in the last decade for cancer treatment. In epigenetic therapy, all efforts are made to modulate gene expression to the normal status. Importantly, recent studies have shown that epigenetic therapy is focusing on the new gene editing technology, CRISPR-Cas9. This tool was found to be able to effectively modulate gene expression and alter almost any sequence in the genome of cells, resulting in events such as a change in acetylation, methylation, or histone modifications. Of note, the CRISPR-Cas9 system can be used for the treatment of cancers caused by epigenetic alterations. The CRISPR-Cas9 system has greater advantages than other available methods, including potent activity, easy design and high velocity as well as the ability to target any DNA or RNA site. In this review, we described epigenetic modulators, which can be used in the CRISPR-Cas9 system, as well as their functions in gene expression alterations that lead to cancer initiation and progression. In addition, we surveyed various species of CRISPR-dead Cas9 (dCas9) systems, a mutant version of Cas9 with no endonuclease activity. Such systems are applicable in epigenetic therapy for gene expression modulation through chemical group editing on nucleosomes and chromatin remodeling, which finally return the cell to the normal status and prevent cancer progression.
PubMed returns only the first 10,000 results — refine the query to see the rest.