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https://www.readbyqxmd.com/read/28938163/tumor-cell-targeted-delivery-of-crispr-cas9-by-aptamer-functionalized-lipopolymer-for-therapeutic-genome-editing-of-vegfa-in-osteosarcoma
#1
Chao Liang, Fangfei Li, Luyao Wang, Zong-Kang Zhang, Chao Wang, Bing He, Jie Li, Zhihao Chen, Atik Badshah Shaikh, Jin Liu, Xiaohao Wu, Songlin Peng, Lei Dang, Baosheng Guo, Xiaojuan He, D W T Au, Cheng Lu, Hailong Zhu, Bao-Ting Zhang, Aiping Lu, Ge Zhang
Osteosarcoma (OS) is a highly aggressive pediatric cancer, characterized by frequent lung metastasis and pathologic bone destruction. Vascular endothelial growth factor A (VEGFA), highly expressed in OS, not only contributes to angiogenesis within the tumor microenvironment via paracrine stimulation of vascular endothelial cells, but also acts as an autocrine survival factor for tumor cell themselves, thus making it a promising therapeutic target for OS. CRISPR/Cas9 is a versatile genome editing technology and holds tremendous promise for cancer treatment...
September 13, 2017: Biomaterials
https://www.readbyqxmd.com/read/28937992/efficient-generation-of-mutations-mediated-by-crispr-cas9-in-the-hairy-root-transformation-system-of-brassica-carinata
#2
Thomas W Kirchner, Markus Niehaus, Thomas Debener, Manfred K Schenk, Marco Herde
A protocol for the induction of site-directed deletions and insertions in the genome of Brassica carinata with CRISPR is described. The construct containing the Cas9 nuclease and the guide RNA (gRNA) was delivered by the hairy root transformation technique, and a successful transformation was monitored by GFP fluorescence. PAGE analysis of an amplified region, presumably containing the deletions and insertions, demonstrated up to seven different indels in one transgenic root and in all analyzed roots a wildtype allele of the modified gene was not detectable...
2017: PloS One
https://www.readbyqxmd.com/read/28934492/concerted-action-of-two-3-cap-independent-translation-enhancers-increases-the-competitive-strength-of-translated-viral-genomes
#3
Zhiyou Du, Olga M Alekhina, Konstantin S Vassilenko, Anne E Simon
Several families of plant viruses evolved cap-independent translation enhancers (3'CITE) in the 3' untranslated regions of their genomic (g)RNAs to compete with ongoing cap-dependent translation of cellular mRNAs. Umbravirus Pea enation mosaic virus (PEMV)2 is the only example where three 3'CITEs enhance translation: the eIF4E-binding Panicum mosaic virus-like translational enhancer (PTE) and ribosome-binding 3' T-shaped structure (TSS) have been found in viruses of different genera, while the ribosome-binding kl-TSS that provides a long-distance interaction with the 5' end is unique...
September 19, 2017: Nucleic Acids Research
https://www.readbyqxmd.com/read/28922955/comparison-of-zfns-versus-crispr-specific-nucleases-for-genome-edition-of-the-wiskott-aldrich-syndrome-locus
#4
Alejandra Gutierrez-Guerrero, Sabina Sanchez-Hernandez, Giuseppe Galvani, Javier Pinedo-Gomez, Almudena Sanchez-Gilabert, Rocio Martin-Guerra, Marien Cobo, Philip Gregory, Michael Holmes, Karim Benabdellah, Francisco Martin
Primary immunodeficiencies (PID), including Wiskott-Aldrich syndrome (WAS), are a main target for genome editing (GE) strategies using specific nucleases (SNs) since a small number of corrected hematopoietic stem cells (HSCs) could cure patients. In this work, we have designed different WAS gene-specific CRISPR/Cas9 systems and compared their efficiency and specificity with homodimeric and heterodimeric WAS-specific Zinc Finger Nucleases (ZFNs) using K562 cells as a cellular model and plasmid nucleofection or integrative-deficient Lentiviral Vectors (IDLVs) for delivery...
September 19, 2017: Human Gene Therapy
https://www.readbyqxmd.com/read/28904745/genome-editing-of-the-hiv-co-receptors-ccr5-and-cxcr4-by-crispr-cas9-protects-cd4-t-cells-from-hiv-1-infection
#5
Zhepeng Liu, Shuliang Chen, Xu Jin, Qiankun Wang, Kongxiang Yang, Chenlin Li, Qiaoqiao Xiao, Panpan Hou, Shuai Liu, Shaoshuai Wu, Wei Hou, Yong Xiong, Chunyan Kong, Xixian Zhao, Li Wu, Chunmei Li, Guihong Sun, Deyin Guo
BACKGROUND: The main approach to treat HIV-1 infection is combination antiretroviral therapy (cART). Although cART is effective in reducing HIV-1 viral load and controlling disease progression, it has many side effects, and is expensive for HIV-1 infected patients who must remain on lifetime treatment. HIV-1 gene therapy has drawn much attention as studies of genome editing tools have progressed. For example, zinc finger nucleases (ZFN), transcription activator like effector nucleases (TALEN) and clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9 have been utilized to successfully disrupt the HIV-1 co-receptors CCR5 or CXCR4, thereby restricting HIV-1 infection...
2017: Cell & Bioscience
https://www.readbyqxmd.com/read/28879858/crispr-mediated-genome-engineering-and-its-application-in-industry
#6
Saeed Kaboli, Hasan Babazada
The CRISPR (clustered regularly interspaced short palindromic repeat)-Cas9 (CRISPR-associated nuclease 9) method has been dramatically changing the field of genome engineering. It is a rapid, highly efficient and versatile tool for precise modification of genome that uses a guide RNA (gRNA) to target Cas9 to a specific sequence. This novel RNA-guided genome-editing technique has become a revolutionary tool in biomedical science and has many innovative applications in different fields. In this review, we briefly introduce the Cas9-mediated genome-editing tool, summarize the recent advances in CRISPR/Cas9 technology to engineer the genomes of a wide variety of organisms, and discuss their applications to treatment of fungal and viral disease...
September 7, 2017: Current Issues in Molecular Biology
https://www.readbyqxmd.com/read/28879857/improving-crispr-cas9-on-target-specificity
#7
Muhammad Jamal, Arif Ullah, Muhammad Ahsan, Rohit Tyagi, Zeshan Habib, Khaista Rehman
The CRISPR-Cas9 has revolutionized the field of molecular biology, medical genetics and medicine. The technology is robust, facile and simple to achieve genome targeting in cells and organisms. However, to propagate these nucleases for therapeutic application, the on-target specificity is of paramount importance. Although the binding and cleavage of off-target sites by Cas9 is issue of concern, however the specificity of CRISPR technology is greatly improved in current research employing the use of engineer nucleases, improved gRNA selection, novel Cas9 orhtologs and the advancement in methods to detect and screen off-target sites and its effects...
September 7, 2017: Current Issues in Molecular Biology
https://www.readbyqxmd.com/read/28868328/creating-a-raw264-7-crispr-cas9-genome-wide-library
#8
Brooke A Napier, Denise M Monack
The bacterial clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9 genome editing tools are used in mammalian cells to knock-out specific genes of interest to elucidate gene function. The CRISPR-Cas9 system requires that the mammalian cell expresses Cas9 endonuclease, guide RNA (gRNA) to lead the endonuclease to the gene of interest, and the PAM sequence that links the Cas9 to the gRNA. CRISPR-Cas9 genome wide libraries are used to screen the effect of each gene in the genome on the cellular phenotype of interest, in an unbiased high-throughput manner...
May 20, 2017: Bio-protocol
https://www.readbyqxmd.com/read/28863137/cas9-grna-targeted-excision-of-cystic-fibrosis-causing-deep-intronic-splicing-mutations-restores-normal-splicing-of-cftr-mrna
#9
David J Sanz, Jennifer A Hollywood, Martina F Scallan, Patrick T Harrison
Cystic Fibrosis is an autosomal recessive disorder caused by mutations in the CFTR gene. CRISPR mediated, template-dependent homology-directed gene editing has been used to correct the most common mutation, c.1521_1523delCTT / p.Phe508del (F508del) which affects ~70% of individuals, but the efficiency was relatively low. Here, we describe a high efficiency strategy for editing of three different rare CFTR mutations which together account for about 3% of individuals with Cystic Fibrosis. The mutations cause aberrant splicing of CFTR mRNA due to the creation of cryptic splice signals that result in the formation of pseudoexons containing premature stop codons c...
2017: PloS One
https://www.readbyqxmd.com/read/28857338/single-nucleotide-editing-without-dna-cleavage-using-crispr-cas9-deaminase-in-the-sea-urchin-embryo
#10
Saba Shevidi, Alicia Uchida, Natalie Schudrowitz, Gary M Wessel, Mamiko Yajima
BACKGROUND: A single base pair mutation in the genome can result in many congenital disorders in humans. The recent gene editing approach using CRISPR/Cas9 has rapidly become a powerful tool to replicate or repair such mutations in the genome. These approaches rely on cleaving DNA, while presenting unexpected risks. RESULTS: In this study, we demonstrate a modified CRISPR/Cas9 system fused to Cytosine DeAminase (Cas9-DA), which induces a single nucleotide conversion in the genome...
August 31, 2017: Developmental Dynamics: An Official Publication of the American Association of Anatomists
https://www.readbyqxmd.com/read/28839466/the-grna-mirna-grna-ternary-cassette-combining-crispr-cas9-with-rnai-approach-strongly-inhibits-hepatitis-b-virus-replication
#11
Jie Wang, Ran Chen, Ruiyang Zhang, Shanlong Ding, Tianying Zhang, Quan Yuan, Guiwen Guan, Xiangmei Chen, Ting Zhang, Hui Zhuang, Frederick Nunes, Timothy Block, Shuang Liu, Zhongping Duan, Ningshao Xia, Zhongwei Xu, Fengmin Lu
The CRISPR/Cas9 system is a novel genome editing technology which has been successfully used to inhibit HBV replication. Here, we described a novel gRNA-microRNA (miRNA)-gRNA ternary cassette driven by a single U6 promoter. With an anti-HBV pri-miR31 mimic integrated between two HBV-specific gRNAs, both gRNAs could be separated from the long transcript of gRNA-miR-HBV-gRNA ternary cassette through Drosha/DGCR8 processing. The results showed that the gRNA-miR-HBV-gRNA ternary cassette could efficiently express two gRNAs and miR-HBV...
2017: Theranostics
https://www.readbyqxmd.com/read/28838224/specific-expression-of-interferon-%C3%AE-induced-by-synergistic-activation-mediator-derived-sam-systems-activates-innate-immunity-and-inhibits-tumorigenesis
#12
Shuai Liu, Xiao Yu, Qiankun Wang, Zhepeng Liu, Qiaoqiao Xiao, Panpan Hou, Ying Hu, Wei Hou, Zhanqiu Yang, Deyin Guo, Shuliang Chen
The synergistic activation mediator (SAM) system can robustly activate endogenous gene expression by a single guide RNA (gRNA). This transcriptional modulation has been shown to enhance gene promoter activity and lead to epigenetic changes. Human interferon-γ is a common natural glycoprotein involved in anti-viral effects and inhibition of cancer cell growth. Large quantities of high-purity interferon-γ are important for medical research and clinical therapy. To investigate the possibility of employing the SAM system to enhance endogenous human interferon-γ with normal function in innate immunity, we designed ten single guide RNAs (sgRNAs) that target 200 bp upstream of the transcription start sites of the interferon-γ genome, which could significantly activate the interferon-γ promoter reporter...
August 25, 2017: Journal of Microbiology and Biotechnology
https://www.readbyqxmd.com/read/28835711/development-of-a-versatile-and-conventional-technique-for-gene-disruption-in-filamentous-fungi-based-on-crispr-cas9-technology
#13
Yan-Mei Zheng, Fu-Long Lin, Hao Gao, Gen Zou, Jiang-Wei Zhang, Gao-Qian Wang, Guo-Dong Chen, Zhi-Hua Zhou, Xin-Sheng Yao, Dan Hu
Filamentous fungi represent an invaluable source of pharmaceutically active compounds. The development of versatile methods to genetically manipulate filamentous fungi is of great value for improving the low yields of bioactive metabolites and expanding chemical diversity. The CRISPR-Cas9-based system has become a common platform for genome editing in a variety of organisms. However, recent application of this technology in filamentous fungi is limited to model strains, a versatile method for efficient gene disruption in different fungi is lacking...
August 23, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28809467/multidimensional-control-of-cas9-by-evolved-rna-polymerase-based-biosensors
#14
Jinyue Pu, Kaitlin Kentala, Bryan C Dickinson
Systems to control Cas9 with spatial and temporal precision offer opportunities to decrease side effects, protect sensitive tissues, and create gene therapies that are only activated at defined times and places. Here, we present the design of new Cas9 controllers based on RNA polymerase (RNAP)-based biosensors that produce gRNAs, thereby regulating target knockout. After development and validation of a new abscisic acid-inducible biosensor to control Cas9, we lowered the background of the system using continuous evolution...
August 15, 2017: ACS Chemical Biology
https://www.readbyqxmd.com/read/28800611/using-local-chromatin-structure-to-improve-crispr-cas9-efficiency-in-zebrafish
#15
Yunru Chen, Shiyang Zeng, Ruikun Hu, Xiangxiu Wang, Weilai Huang, Jiangfang Liu, Luying Wang, Guifen Liu, Ying Cao, Yong Zhang
Although the CRISPR/Cas9 has been successfully applied in zebrafish, considerable variations in efficiency have been observed for different gRNAs. The workload and cost of zebrafish mutant screening is largely dependent on the mutation rate of injected embryos; therefore, selecting more effective gRNAs is especially important for zebrafish mutant construction. Besides the sequence features, local chromatin structures may have effects on CRISPR/Cas9 efficiency, which remain largely unexplored. In the only related study in zebrafish, nucleosome organization was not found to have an effect on CRISPR/Cas9 efficiency, which is inconsistent with recent studies in vitro and in mammalian cell lines...
2017: PloS One
https://www.readbyqxmd.com/read/28800587/highly-efficient-gene-inactivation-by-adenoviral-crispr-cas9-in-human-primary-cells
#16
Olaf Voets, Frans Tielen, Edo Elstak, Julian Benschop, Max Grimbergen, Jan Stallen, Richard Janssen, Andre van Marle, Christian Essrich
Phenotypic assays using human primary cells are highly valuable tools for target discovery and validation in drug discovery. Expression knockdown (KD) of such targets in these assays allows the investigation of their role in models of disease processes. Therefore, efficient and fast modes of protein KD in phenotypic assays are required. The CRISPR/Cas9 system has been shown to be a versatile and efficient means of gene inactivation in immortalized cell lines. Here we describe the use of adenoviral (AdV) CRISPR/Cas9 vectors for efficient gene inactivation in two human primary cell types, normal human lung fibroblasts and human bronchial epithelial cells...
2017: PloS One
https://www.readbyqxmd.com/read/28794219/dodging-silver-bullets-good-crispr-gene-drive-design-is-critical-for-eradicating-exotic-vertebrates
#17
Thomas A A Prowse, Phillip Cassey, Joshua V Ross, Chandran Pfitzner, Talia A Wittmann, Paul Thomas
Self-replicating gene drives that can spread deleterious alleles through animal populations have been promoted as a much needed but controversial 'silver bullet' for controlling invasive alien species. Homing-based drives comprise an endonuclease and a guide RNA (gRNA) that are replicated during meiosis via homologous recombination. However, their efficacy for controlling wild populations is threatened by inherent polymorphic resistance and the creation of resistance alleles via non-homologous end-joining (NHEJ)-mediated DNA repair...
August 16, 2017: Proceedings. Biological Sciences
https://www.readbyqxmd.com/read/28790199/in-vivo-genome-editing-restores-dystrophin-expression-and-cardiac-function-in-dystrophic-mice
#18
Mona El Refaey, Li Xu, Yandi Gao, Benjamin D Canan, Tm A Adesanya, Sarah C Warner, Keiko Akagi, David E Symer, Peter J Mohler, Jianjie Ma, Paul M Janssen, Renzhi Han
Rationale: Duchenne muscular dystrophy (DMD) is a severe inherited form of muscular dystrophy caused by mutations in the reading frame of the dystrophin gene disrupting its protein expression. Dystrophic cardiomyopathy is a leading cause of death in DMD patients and currently no effective treatment exists to halt its progression. Recent advancement in genome editing technologies offers a promising therapeutic approach in restoring dystrophin protein expression. However, the impact of this approach on DMD cardiac function has yet to be evaluated...
August 8, 2017: Circulation Research
https://www.readbyqxmd.com/read/28757163/evaluating-synthetic-activation-and-repression-of-neuropsychiatric-related-genes-in-hipsc-derived-npcs-neurons-and-astrocytes
#19
Seok-Man Ho, Brigham J Hartley, Erin Flaherty, Prashanth Rajarajan, Rawan Abdelaal, Ifeanyi Obiorah, Natalie Barretto, Hamza Muhammad, Hemali P Phatnani, Schahram Akbarian, Kristen J Brennand
Modulation of transcription, either synthetic activation or repression, via dCas9-fusion proteins is a relatively new methodology with the potential to facilitate high-throughput up- or downregulation studies of gene function. Genetic studies of neurodevelopmental disorders have identified a growing list of risk variants, including both common single-nucleotide variants and rare copy-number variations, many of which are associated with genes having limited functional annotations. By applying a CRISPR-mediated gene-activation/repression platform to populations of human-induced pluripotent stem cell-derived neural progenitor cells, neurons, and astrocytes, we demonstrate that it is possible to manipulate endogenous expression levels of candidate neuropsychiatric risk genes across these three cell types...
August 8, 2017: Stem Cell Reports
https://www.readbyqxmd.com/read/28750323/differential-use-of-3-cites-by-the-subgenomic-rna-of-pea-enation-mosaic-virus-2
#20
Feng Gao, Anne E Simon
The genomic RNA (gRNA) of Pea enation mosaic virus 2 (PEMV2) is the template for p33 and -1 frameshift product p94. The PEMV2 subgenomic RNA (sgRNA) encodes two overlapping ORFs, p26 and p27, which are required for movement and stability of the gRNA. Efficient translation of p33 requires two of three 3' proximal cap-independent translation enhancers (3'CITEs): the kl-TSS, which binds ribosomes and engages in a long-distance interaction with the 5'end; and the adjacent eIF4E-binding PTE. Unlike the gRNA, all three 3'CITEs were required for efficient translation of the sgRNA, which included the ribosome-binding 3'TSS...
October 2017: Virology
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