3,635 results match your criteria crispr-associated protein

Progress of genome editing technology and developmental biology useful for radiation research.

J Radiat Res 2021 May;62(Supplement_1):i53-i63

Department of Disease Model, Research Institute of Radiation Biology and Medicine, Hiroshima University, Hiroshima 734-8553, Japan.

Following the development of genome editing technology, it has become more feasible to create genetically modified animals such as knockout (KO), knock-in, and point-mutated animals. The genome-edited animals are useful to investigate the roles of various functional genes in many fields of biological science including radiation research. Nevertheless, some researchers may experience difficulty in generating genome-edited animals, probably due to the requirement for equipment and techniques for embryo manipulation and handling. Read More

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[Structure-based optimization and design of CRISPR protein xCas9].

Sheng Wu Gong Cheng Xue Bao 2021 Apr;37(4):1385-1395

School of Life Sciences, Fudan University, Shanghai 200438, China.

Streptococcus pyogenes Cas9 (SpCas9) has become a powerful genome editing tool, but has a limited range of recognizable protospacer adjacent motifs (PAMs) and shows off-target effects. To address these issues, we present a rational approach to optimize the xCas9 mutant derived from SpCas9 by directed evolution. Firstly, energy minimization with the Rosetta program was applied to optimize the three-dimensional structure of Cas9 to obtain the lowest energy conformation. Read More

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SEMMs: Somatically Engineered Mouse Models. A New Tool for Disease Modeling for Basic and Translational Research.

Front Oncol 2021 23;11:667189. Epub 2021 Apr 23.

Department of Translational Oncology, Genentech, Inc., South San Francisco, CA, United States.

Most experimental oncology therapies fail during clinical development despite years of preclinical testing rationalizing their use. This begs the question of whether the current preclinical models used for evaluating oncology therapies adequately capture patient heterogeneity and response to therapy. Most of the preclinical work is based on xenograft models where tumor mis-location and the lack of the immune system represent a major limitation for the translatability of many observations from preclinical models to patients. Read More

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CRISPR-Cas-mediated chromosome engineering for crop improvement and synthetic biology.

Nat Plants 2021 May 6. Epub 2021 May 6.

Botanical Institute, Karlsruhe Institute of Technology, Karlsruhe, Germany.

Plant breeding relies on the presence of genetic variation, as well as on the ability to break or stabilize genetic linkages between traits. The development of the genome-editing tool clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR-associated protein (Cas) has allowed breeders to induce genetic variability in a controlled and site-specific manner, and to improve traits with high efficiency. However, the presence of genetic linkages is a major obstacle to the transfer of desirable traits from wild species to their cultivated relatives. Read More

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The effect of CRISPR constructs microinjection on the expression of developmental genes in Rag1 knocked-out mice embryo.

Vet Med Sci 2021 May 6. Epub 2021 May 6.

Department of Gametes and Cloning, Research Institute of Animal Embryo Technology, Shahrekord University, Shahrekord, Iran.

Despite all the advances in the production of transgenic mice, the production efficiency of these animal models is still low. Given that the expression of developmental genes has a critical role in growth and development of embryo, we determined the expression pattern of pluripotency, trophectoderm and imprinting genes in the Rag1 (recombination-activating gene 1) knocked-out blastocysts resulting from microinjection of CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9) constructs into the zygote cytoplasm of C57bl6 mice. Following microinjection, the embryos were cultured and the gene expression of developed blastocysts and natural blastocysts (Sham and control groups) were evaluated using real-time PCR. Read More

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CRISPR-Cas technology based genome editing for modification of salinity stress tolerance responses in rice (Oryza sativa L.).

Mol Biol Rep 2021 May 5. Epub 2021 May 5.

Department of Bioinformatics and Biosciences, Capital University of Science and Technology, Islamabad, Pakistan.

Clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR associated protein (Cas) technology is an effective tool for site-specific genome editing, used to precisely induce mutagenesis in different plant species including rice. Salinity is one of the most stressful environmental constraints affecting agricultural productivity worldwide. As plant adaptation to salinity stress is under polygenic control therefore, 51 rice genes have been identified that play crucial role in response to salinity. Read More

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A novel non-viral delivery method that enables efficient engineering of primary human T cells for ex vivo cell therapy applications.

Cytotherapy 2021 Apr 30. Epub 2021 Apr 30.

Avectas Ltd, Maynooth, Ireland. Electronic address:

Background Aims: Next-generation immune cell therapy products will require complex modifications using engineering technologies that can maintain high levels of cell functionality. Non-viral engineering methods have the potential to address limitations associated with viral vectors. However, while electroporation is the most widely used non-viral modality, concerns about its effects on cell functionality have led to the exploration of alternative approaches. Read More

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Patient-Specific induced Pluripotent Stem Cell-Derived Hepatocyte-Like Cells as a Model to Study Autosomal Recessive Hypercholesterolemia.

Stem Cells Dev 2021 May 2. Epub 2021 May 2.

Associatie KU Leuven, 418666, Department of Development and Regeneration, Stem Cell Biology and Embryology, Interdepartmental Stem Cell Institute, Leuven, Belgium;

Autosomal recessive hypercholesterolemia (ARH) is a rare monogenic disorder caused by pathogenic variants in the low-density lipoprotein receptor (LDLR) adaptor protein-1 (LDLRAP1) gene, encoding for the LDLRAP1 protein, which impairs internalization of hepatic LDLR. There are variable responses of ARH patients to treatment and the pathophysiological mechanism(s) for this variability remains unclear. This is in part caused by absence of reliable cellular models to evaluate the effect of LDLRAP1 mutations on LDLRAP1 protein function and its role in LDLR internalization. Read More

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CRISPR-Cas systems: Challenges and future prospects.

Prog Mol Biol Transl Sci 2021 15;180:141-151. Epub 2021 Feb 15.

Department of Biosciences, School of Science, Indrashil University, Rajpur, Mehsana, Gujarat, India. Electronic address:

The advancement gained over the past couple of decades in clustered regularly interspaced short palindromic repeats and CRISPR associated proteins (CRISPR-Cas) systems have revolutionized the field of synthetic biology, therapeutics, diagnostics and metabolic engineering. The technique has enabled the process of genome editing to be very precise, rapid, cost-effective and highly efficient which were the downfalls for the previously debuted zinc finger nucleases (ZFN) and transcription activator-like effector nucleases (TALEN) technologies. However, despite its great potential, challenges including off-target activity, method of delivery, ethical and regulatory issues still remain unresolved for the CRISPR-Cas systems. Read More

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February 2021

A Simple and Quick Method for Loading Proteins in Extracellular Vesicles.

Pharmaceuticals (Basel) 2021 Apr 13;14(4). Epub 2021 Apr 13.

Department of Biochemistry and Molecular Biology, Mayo Clinic, Jacksonville, FL 32224, USA.

Extracellular vesicles (EVs) mediate intercellular transport of biomolecular cargo in the body, making them promising delivery vehicles for bioactive compounds. Genetic engineering of producer cells has enabled encapsulation of therapeutic proteins in EVs. However, genetic engineering approaches can be expensive, time-consuming, and incompatible with certain EV sources, such as human plasma and bovine milk. Read More

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Characterisation of Phage Susceptibility Variation in Serovar Typhimurium DT104 and DT104b.

Microorganisms 2021 Apr 17;9(4). Epub 2021 Apr 17.

School of Life Sciences, College of Liberal Arts and Sciences, University of Westminster, 115 New Cavendish Street, Fitzrovia, London W1W 6XH, UK.

The surge in mortality and morbidity rates caused by multidrug-resistant (MDR) bacteria prompted a renewal of interest in bacteriophages (phages) as clinical therapeutics and natural biocontrol agents. Nevertheless, bacteria and phages are continually under the pressure of the evolutionary phage-host arms race for survival, which is mediated by co-evolving resistance mechanisms. In Anderson phage typing scheme of Typhimurium, the epidemiologically related definitive phage types, DT104 and DT104b, display significantly different phage susceptibility profiles. Read More

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Improved CRISPR/Cas9 Tools for the Rapid Metabolic Engineering of .

Int J Mol Sci 2021 Apr 2;22(7). Epub 2021 Apr 2.

INP, TBI, INSA, UPS, Université de Toulouse, 31400 Toulouse, France.

Clustered regularly interspaced short palindromic repeats (CRISPR)/Cas (CRISPR-associated proteins)9 tools have revolutionized biology-several highly efficient tools have been constructed that have resulted in the ability to quickly engineer model bacteria, for example, . However, the use of CRISPR/Cas9 tools has lagged behind in non-model bacteria, hampering engineering efforts. Here, we developed improved CRISPR/Cas9 tools to enable efficient rapid metabolic engineering of the industrially relevant bacterium . Read More

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V-Set and Immunoglobulin Domain-Containing 1 (VSIG1), Predominantly Expressed in Testicular Germ Cells, Is Dispensable for Spermatogenesis and Male Fertility in Mice.

Animals (Basel) 2021 Apr 7;11(4). Epub 2021 Apr 7.

College of Pharmacy, Catholic University of Daegu, Gyeongsan-si 38430, Korea.

To elucidate the functional role of V-set and immunoglobulin domain-containing 1 (VSIG1) in spermatogenesis and fertilization, we knocked out (KO) VSIG1 in a mouse embryo using CRISPR/Cas9 (Clustered regularly interspaced short palindromic repeat/CRISPR-associated protein 9) -mediated genome editing. Reverse transcription PCR was performed using cDNA synthesized from VSIG1 KO testis RNA. Although Western blot analysis using a specific antibody to VSIG1 confirmed VSIG1 protein defects in the KO mice, hematoxylin-eosin staining analysis was similar in the KO and wild-type mice. Read More

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CRISPR-Cas tools to study gene function in cytokinesis.

J Cell Sci 2021 Apr 22;134(8). Epub 2021 Apr 22.

Biology Department, Concordia University, Montréal, QC, H4B 1R6, Canada.

Cytokinesis is the process that separates a cell into two daughter cells at the end of mitosis. Most of our knowledge of cytokinesis comes from overexpression studies, which affects our interpretation of protein function. Gene editing can circumvent this issue by introducing functional mutations or fluorescent probes directly into a gene locus. Read More

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Kindlin2 regulates neural crest specification via integrin-independent regulation of the FGF signaling pathway.

Development 2021 Apr 28. Epub 2021 Apr 28.

Key Laboratory for Regenerative Medicine, Ministry of Education, School of Biomedical Sciences, Faculty of Medicine, The Chinese University of Hong Kong, Hong Kong SAR, China.

The focal adhesion protein Kindlin2 is essential for integrin activation, a process that is fundamental to cell-extracellular matrix adhesion. Kindlin2 is widely expressed in mouse embryos, and its absence causes lethality at the peri-implantation stage due to the failure to trigger integrin activation. The function of kindlin2 during embryogenesis has not yet been fully elucidated as a result of this early embryonic lethality. Read More

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[Rapid screening of single guide RNA targeting pig genome and the harvesting of monoclonal cells by microarray seal].

Sheng Wu Yi Xue Gong Cheng Xue Za Zhi 2021 Feb;38(1):111-121

Department of Pathology, West China Hospital, Sichuan University, Chengdu 610041, P.R.China;Institute of Clinical Pathology, West China Hospital, Sichuan University, Chengdu 610041, P.R.China.

The emergence of regular short repetitive palindromic sequence clusters (CRISPR) and CRISPR- associated proteins 9 (Cas9) gene editing technology has greatly promoted the wide application of genetically modified pigs. Efficient single guide RNA (sgRNA) is the key to the success of gene editing using CRISPR/Cas9 technology. For large animals with a long reproductive cycle, such as pigs, it is necessary to screen out efficient sgRNA to avoid wasting time and resource costs before animal experiments. Read More

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February 2021

Proteome and Phosphoproteome Analyses Reveal the Kinase Regulatory Network Involved in Glycogen Synthesis Kinase 3β.

Front Genet 2021 7;12:657140. Epub 2021 Apr 7.

The First Affiliated Hospital of Zhengzhou University, Zhengzhou, China.

Diabetic nephropathy is the most common chronic kidney disease in the world and the main cause of end-stage renal disease (ESRD). The structural integrity of podocytes is fundamental to the normal function of the glomerulus, and the role of glycogen synthase kinase 3β (GSK-3β) in podocytes is complicated. A thorough understanding of GSK-3β is crucial to understand the mechanism of diabetic nephropathy. Read More

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DNA Repair Pathway Choices in CRISPR-Cas9-Mediated Genome Editing.

Trends Genet 2021 Apr 22. Epub 2021 Apr 22.

Department of Biochemistry and Molecular Biophysics, Columbia University, New York, NY 10032, USA. Electronic address:

Many clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR-associated protein 9 (Cas9)-based genome editing technologies take advantage of Cas nucleases to induce DNA double-strand breaks (DSBs) at desired locations within a genome. Further processing of the DSBs by the cellular DSB repair machinery is then necessary to introduce desired mutations, sequence insertions, or gene deletions. Thus, the accuracy and efficiency of genome editing are influenced by the cellular DSB repair pathways. Read More

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CRISPR-based genome editing in primary human pancreatic islet cells.

Nat Commun 2021 04 23;12(1):2397. Epub 2021 Apr 23.

Department of Developmental Biology, Stanford University School of Medicine, Stanford, CA, USA.

Gene targeting studies in primary human islets could advance our understanding of mechanisms driving diabetes pathogenesis. Here, we demonstrate successful genome editing in primary human islets using clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated protein 9 (Cas9). CRISPR-based targeting efficiently mutated protein-coding exons, resulting in acute loss of islet β-cell regulators, like the transcription factor PDX1 and the K channel subunit KIR6. Read More

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Activated IL-6 signaling contributes to the pathogenesis of, and is a novel therapeutic target for, CALR-mutated MPNs.

Blood Adv 2021 Apr;5(8):2184-2195

Center of Research and Innovation of Myeloproliferative Neoplasms (CRIMM), Department of Experimental and Clinical Medicine, University of Florence, Florence, Italy.

Calreticulin (CALR), an endoplasmic reticulum-associated chaperone, is frequently mutated in myeloproliferative neoplasms (MPNs). Mutated CALR promotes downstream JAK2/STAT5 signaling through interaction with, and activation of, the thrombopoietin receptor (MPL). Here, we provide evidence of a novel mechanism contributing to CALR-mutated MPNs, represented by abnormal activation of the interleukin 6 (IL-6)-signaling pathway. Read More

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Multiplex Genome Editing in Chinese Hamster Ovary Cell Line Using All-in-One and HITI CRISPR Technology.

Adv Pharm Bull 2021 Feb 15;11(2):343-350. Epub 2020 Apr 15.

Department of Physiology & Biophysics, School of Medicine, Case Western Reserve University, Ohio, USA.

CRISPR/Cas9 gene editing technology has revolutionized gene manipulation by providing the opportunity of gene knock out/in, transcriptional modification and base editing. The application of this system extended into different eras of biology, from cell development to animal modeling. Various generations of CRISPR technology have been developed to make genome editing easy which resulted in rapid protocols for amelioration of a large genome. Read More

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February 2021

CRISPR/Cas12a-based on-site diagnostics of Cryptosporidium parvum IId-subtype-family from human and cattle fecal samples.

Parasit Vectors 2021 Apr 20;14(1):208. Epub 2021 Apr 20.

College of Animal Science and Veterinary Medicine, Longzihu Campus of Henan Agricultural University, No. 15 Longzihu University Area, Zhengzhou New District, Zhengzhou, 450046, People's Republic of China.

Background: Cryptosporidium parvum is an enteric protozoan parasite with zoonotic importance and can cause cryptosporidiosis in humans as well as domestic and wild animals worldwide. The IId subtype family (SF) is one of the most prevalent subtypes of C. parvum. Read More

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Delivery of the Cas9/sgRNA Ribonucleoprotein Complex in Immortalized and Primary Cells via Virus-like Particles ("Nanoblades").

J Vis Exp 2021 03 31(169). Epub 2021 Mar 31.

LBMC, Laboratoire de Biologie et Modélisation de la Cellule Univ Lyon, ENS de Lyon, Université Claude Bernard Lyon 1, CNRS, UMR 5239, INSERM, U1210, Lyon, 69007, France;

The clustered regularly interspaced short palindromic repeats (CRISPR)-Cas system has democratized genome-editing in eukaryotic cells and led to the development of numerous innovative applications. However, delivery of the Cas9 protein and single-guide RNA (sgRNA) into target cells can be technically challenge. Classical viral vectors, such as those derived from lentiviruses (LVs) or adeno-associated viruses (AAVs), allow for efficient delivery of transgenes coding for the Cas9 protein and its associated sgRNA in many primary cells and in vivo. Read More

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Gene Silencing Through CRISPR Interference in Bacteria: Current Advances and Future Prospects.

Front Microbiol 2021 31;12:635227. Epub 2021 Mar 31.

State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, China.

Functional genetic screening is an important method that has been widely used to explore the biological processes and functional annotation of genetic elements. CRISPR/Cas (Clustered regularly interspaced short palindromic repeat sequences/CRISPR-associated protein) is the newest tool in the geneticist's toolbox, allowing researchers to edit a genome with unprecedented ease, accuracy, and high-throughput. Most recently, CRISPR interference (CRISPRi) has been developed as an emerging technology that exploits the catalytically inactive Cas9 (dCas9) and single-guide RNA (sgRNA) to repress sequence-specific genes. Read More

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Structure-guided engineering of adenine base editor with minimized RNA off-targeting activity.

Nat Commun 2021 04 16;12(1):2287. Epub 2021 Apr 16.

Precise Genome Engineering Center, School of Life Sciences, Guangzhou University, Guangzhou, China.

Both adenine base editors (ABEs) and cytosine base editors (CBEs) have been recently revealed to induce transcriptome-wide RNA off-target editing in a guide RNA-independent manner. Here we construct a reporter system containing E.coli Hokb gene with a tRNA-like motif for robust detection of RNA editing activities as the optimized ABE, ABEmax, induces highly efficient A-to-I (inosine) editing within an E. Read More

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Methods and Techniques to Facilitate the Development of NT as an Effective, Therapeutic Oncolytic Bacteria.

Front Microbiol 2021 29;12:624618. Epub 2021 Mar 29.

Cell and Molecular Biology Program, North Dakota State University, Fargo, ND, United States.

The tumor microenvironment is characterized by anomalous vascularization, hypoxia, and acidity at the core of solid tumors that culminates in concentrated necrosis and immune system dysregulation among other effects. While this environment presents several challenges for the development of oncotherapeutics that deliver their activity via the enhanced permeability and retention (EPR) effect of the leaky blood vessels around a tumor, oncolytic bacteria, or a class of bacteria with a noted capacity to lyse solid tumors, are attracted to the very environment found at the center of solid tumors that confounds other therapeutics. It is this capacity that allows for a potent, active penetration from the tumor margins into the core, and subsequent colonization to facilitate lysis and immune reactivation. Read More

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Learning cis-regulatory principles of ADAR-based RNA editing from CRISPR-mediated mutagenesis.

Nat Commun 2021 04 12;12(1):2165. Epub 2021 Apr 12.

Department of Genetics, Stanford University, Stanford, CA, USA.

Adenosine-to-inosine (A-to-I) RNA editing catalyzed by ADAR enzymes occurs in double-stranded RNAs. Despite a compelling need towards predictive understanding of natural and engineered editing events, how the RNA sequence and structure determine the editing efficiency and specificity (i.e. Read More

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In vivo genome editing in mouse restores dystrophin expression in Duchenne muscular dystrophy patient muscle fibers.

Genome Med 2021 Apr 12;13(1):57. Epub 2021 Apr 12.

CAS Key Laboratory of Regenerative Biology, Guangdong Provincial Key Laboratory of Stem Cell and Regenerative Medicine, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou, 510530, China.

Background: Mutations in the DMD gene encoding dystrophin-a critical structural element in muscle cells-cause Duchenne muscular dystrophy (DMD), which is the most common fatal genetic disease. Clustered regularly interspaced short palindromic repeat (CRISPR)-mediated gene editing is a promising strategy for permanently curing DMD.

Methods: In this study, we developed a novel strategy for reframing DMD mutations via CRISPR-mediated large-scale excision of exons 46-54. Read More

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Current status of the application of gene editing in pigs.

J Reprod Dev 2021 Apr 10. Epub 2021 Apr 10.

Faculty of Bioscience and Bioindustry, Tokushima University, Tokushima 770-8513, Japan.

Genetically modified animals, especially rodents, are widely used in biomedical research. However, non-rodent models are required for efficient translational medicine and preclinical studies. Owing to the similarity in the physiological traits of pigs and humans, genetically modified pigs may be a valuable resource for biomedical research. Read More

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