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CRISPR AND Zebrafish

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https://www.readbyqxmd.com/read/28815495/ligand-controlled-site-specific-recombination-in-zebrafish
#1
Avinash Chekuru, Veronika Kuscha, Stefan Hans, Michael Brand
Cre-mediated site-specific recombination has emerged as an indispensable tool for the precise manipulation of genomes allowing lineage-tracing studies, temporal and spatial misexpressions, and in particular the generation of conditional knockout alleles. Previously, we and others showed that Cre and its ligand-inducible variant CreER(T2) are also highly efficient in the developing and adult zebrafish. The number of Cre driver and effector lines is currently still limited in zebrafish. However, the recent advent of novel genome editing tools such as TALEN and CRISPR/Cas will significantly increase interest in the conditional Cre/lox-technology in this organism...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28805828/mutations-in-keops-complex-genes-cause-nephrotic-syndrome-with-primary-microcephaly
#2
Daniela A Braun, Jia Rao, Geraldine Mollet, David Schapiro, Marie-Claire Daugeron, Weizhen Tan, Olivier Gribouval, Olivia Boyer, Patrick Revy, Tilman Jobst-Schwan, Johanna Magdalena Schmidt, Jennifer A Lawson, Denny Schanze, Shazia Ashraf, Jeremy F P Ullmann, Charlotte A Hoogstraten, Nathalie Boddaert, Bruno Collinet, Gaëlle Martin, Dominique Liger, Svjetlana Lovric, Monica Furlano, I Chiara Guerrera, Oraly Sanchez-Ferras, Jennifer F Hu, Anne-Claire Boschat, Sylvia Sanquer, Björn Menten, Sarah Vergult, Nina De Rocker, Merlin Airik, Tobias Hermle, Shirlee Shril, Eugen Widmeier, Heon Yung Gee, Won-Il Choi, Carolin E Sadowski, Werner L Pabst, Jillian K Warejko, Ankana Daga, Tamara Basta, Verena Matejas, Karin Scharmann, Sandra D Kienast, Babak Behnam, Brendan Beeson, Amber Begtrup, Malcolm Bruce, Gaik-Siew Ch'ng, Shuan-Pei Lin, Jui-Hsing Chang, Chao-Huei Chen, Megan T Cho, Patrick M Gaffney, Patrick E Gipson, Chyong-Hsin Hsu, Jameela A Kari, Yu-Yuan Ke, Cathy Kiraly-Borri, Wai-Ming Lai, Emmanuelle Lemyre, Rebecca Okashah Littlejohn, Amira Masri, Mastaneh Moghtaderi, Kazuyuki Nakamura, Fatih Ozaltin, Marleen Praet, Chitra Prasad, Agnieszka Prytula, Elizabeth R Roeder, Patrick Rump, Rhonda E Schnur, Takashi Shiihara, Manish D Sinha, Neveen A Soliman, Kenza Soulami, David A Sweetser, Wen-Hui Tsai, Jeng-Daw Tsai, Rezan Topaloglu, Udo Vester, David H Viskochil, Nithiwat Vatanavicharn, Jessica L Waxler, Klaas J Wierenga, Matthias T F Wolf, Sik-Nin Wong, Sebastian A Leidel, Gessica Truglio, Peter C Dedon, Annapurna Poduri, Shrikant Mane, Richard P Lifton, Maxime Bouchard, Peter Kannu, David Chitayat, Daniella Magen, Bert Callewaert, Herman van Tilbeurgh, Martin Zenker, Corinne Antignac, Friedhelm Hildebrandt
Galloway-Mowat syndrome (GAMOS) is an autosomal-recessive disease characterized by the combination of early-onset nephrotic syndrome (SRNS) and microcephaly with brain anomalies. Here we identified recessive mutations in OSGEP, TP53RK, TPRKB, and LAGE3, genes encoding the four subunits of the KEOPS complex, in 37 individuals from 32 families with GAMOS. CRISPR-Cas9 knockout in zebrafish and mice recapitulated the human phenotype of primary microcephaly and resulted in early lethality. Knockdown of OSGEP, TP53RK, or TPRKB inhibited cell proliferation, which human mutations did not rescue...
August 14, 2017: Nature Genetics
https://www.readbyqxmd.com/read/28800611/using-local-chromatin-structure-to-improve-crispr-cas9-efficiency-in-zebrafish
#3
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/28795449/zebrafish-models-of-orofacial-clefts
#4
REVIEW
Kaylia Duncan, Kusumika Mukherjee, Robert A Cornell, Eric C Liao
Zebrafish is a model organism that affords experimental advantages toward investigating the normal function of genes associated with congenital birth defects. Here we summarize zebrafish studies of genes implicated in orofacial cleft (OFC). The most common use of zebrafish in this context has been to explore the normal function an OFC-associated gene product in craniofacial morphogenesis by inhibiting expression of its zebrafish ortholog. The most frequently deployed method has been to inject embryos with antisense morpholino oligonucleotides targeting the desired transcript...
August 10, 2017: Developmental Dynamics: An Official Publication of the American Association of Anatomists
https://www.readbyqxmd.com/read/28767718/zygotic-vinculin-is-not-essential-for-embryonic-development-in-zebrafish
#5
Mitchell K L Han, Gerard N M van der Krogt, Johan de Rooij
The formation of multicellular tissues during development is governed by mechanical forces that drive cell shape and tissue architecture. Protein complexes at sites of adhesion to the extracellular matrix (ECM) and cell neighbors, not only transmit these mechanical forces, but also allow cells to respond to changes in force by inducing biochemical feedback pathways. Such force-induced signaling processes are termed mechanotransduction. Vinculin is a central protein in mechanotransduction that in both integrin-mediated cell-ECM and cadherin-mediated cell-cell adhesions mediates force-induced cytoskeletal remodeling and adhesion strengthening...
2017: PloS One
https://www.readbyqxmd.com/read/28767326/tracking-of-indels-by-decomposition-is-a-simple-and-effective-method-to-assess-efficiency-of-guide-rnas-in-zebrafish
#6
Christelle Etard, Swarnima Joshi, Johannes Stegmaier, Ralf Mikut, Uwe Strähle
A bottleneck in CRISPR/Cas9 genome editing is variable efficiencies of in silico-designed gRNAs. We evaluated the sensitivity of the TIDE method (Tracking of Indels by DEcomposition) introduced by Brinkman et al. in 2014 for assessing the cutting efficiencies of gRNAs in zebrafish. We show that this simple method, which involves bulk polymerase chain reaction amplification and Sanger sequencing, is highly effective in tracking well-performing gRNAs in pools of genomic DNA derived from injected embryos. The method is equally effective for tracing INDELs in heterozygotes...
August 2, 2017: Zebrafish
https://www.readbyqxmd.com/read/28767277/using-zebrafish-to-test-the-genetic-basis-of-human-craniofacial-diseases
#7
R Grecco Machado, B Frank Eames
Genome-wide association studies (GWASs) opened an innovative and productive avenue to investigate the molecular basis of human craniofacial disease. However, GWASs identify candidate genes only; they do not prove that any particular one is the functional villain underlying disease or just an unlucky genomic bystander. Genetic manipulation of animal models is the best approach to reveal which genetic loci identified from human GWASs are functionally related to specific diseases. The purpose of this review is to discuss the potential of zebrafish to resolve which candidate genetic loci are mechanistic drivers of craniofacial diseases...
August 1, 2017: Journal of Dental Research
https://www.readbyqxmd.com/read/28751305/dlx3b-4b-is-required-for-early-born-but-not-later-forming-sensory-hair-cells-during-zebrafish-inner-ear-development
#8
Simone Schwarzer, Sandra Spieß, Michael Brand, Stefan Hans
Morpholino-mediated knockdown has shown that the homeodomain transcription factors Dlx3b and Dlx4b are essential for proper induction of the otic-epibranchial progenitor domain (OEPD) as well as subsequent formation of sensory hair cells in the developing zebrafish inner ear. However, increasing use of reverse genetic approaches has revealed poor correlation between morpholino-induced and mutant phenotypes. Using CRISPR/Cas9-mediated mutagenesis, we generated a defined deletion eliminating the entire open reading frames of dlx3b and dlx4b (dlx3b/4b) and investigated a potential phenotypic difference between mutants and morpholino-mediated knockdown...
July 27, 2017: Biology Open
https://www.readbyqxmd.com/read/28747644/active-nuclear-transcriptome-analysis-reveals-inflammasome-dependent-mechanism-for-early-neutrophil-response-to-mycobacterium-marinum
#9
Amy Kenyon, Daria Gavriouchkina, Jernej Zorman, Giorgio Napolitani, Vincenzo Cerundolo, Tatjana Sauka-Spengler
The mechanisms governing neutrophil response to Mycobacterium tuberculosis remain poorly understood. In this study we utilise biotagging, a novel genome-wide profiling approach based on cell type-specific in vivo biotinylation in zebrafish to analyse the initial response of neutrophils to Mycobacterium marinum, a close genetic relative of M. tuberculosis used to model tuberculosis. Differential expression analysis following nuclear RNA-seq of neutrophil active transcriptomes reveals a significant upregulation in both damage-sensing and effector components of the inflammasome, including caspase b, NLRC3 ortholog (wu: fb15h11) and il1β...
July 26, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28740134/programmable-base-editing-of-zebrafish-genome-using-a-modified-crispr-cas9-system
#10
Yihan Zhang, Wei Qin, Xiaochan Lu, Jason Xu, Haigen Huang, Haipeng Bai, Song Li, Shuo Lin
Precise genetic modifications in model animals are essential for biomedical research. Here, we report a programmable "base editing" system to induce precise base conversion with high efficiency in zebrafish. Using cytidine deaminase fused to Cas9 nickase, up to 28% of site-specific single-base mutations are achieved in multiple gene loci. In addition, an engineered Cas9-VQR variant with 5'-NGA PAM specificities is used to induce base conversion in zebrafish. This shows that Cas9 variants can be used to expand the utility of this technology...
July 25, 2017: Nature Communications
https://www.readbyqxmd.com/read/28739660/a-gene-implicated-in-activation-of-retinoic-acid-receptor-targets-is-a-novel-renal-agenesis-gene-in-humans
#11
Patrick D Brophy, Maria Rasmussen, Mrutyunjaya Parida, Greg Bonde, Benjamin W Darbro, Xiaojing Hong, Jason C Clarke, Kevin A Peterson, James Denegre, Michael Schneider, Caroline R Sussman, Lone Sunde, Dorte L Lildballe, Jens Michael Hertz, Robert A Cornell, Stephen A Murray, John R Manak
Renal agenesis (RA) is one of the more extreme examples of congenital anomalies of the kidney and urinary tract (CAKUT). Bilateral renal agenesis is almost invariably fatal at birth, and unilateral renal agenesis can lead to future health issues including end stage renal disease. Genetic investigations have identified several gene variants which cause RA, including EYA1, LHX1, and WT1 However, whereas compound null mutations of genes encoding α and γ retinoic acid receptors (RARs) cause RA in mice, to date there have been no reports of variants in RAR genes causing RA in humans...
July 24, 2017: Genetics
https://www.readbyqxmd.com/read/28718799/zebrafish-as-a-model-organism-for-the-development-of-drugs-for-skin-cancer
#12
REVIEW
Fatemeh Bootorabi, Hamed Manouchehri, Reza Changizi, Harlan Barker, Elisabetta Palazzo, Annalisa Saltari, Mataleena Parikka, Carlo Pincelli, Ashok Aspatwar
Skin cancer, which includes melanoma and squamous cell carcinoma, represents the most common type of cutaneous malignancy worldwide, and its incidence is expected to rise in the near future. This condition derives from acquired genetic dysregulation of signaling pathways involved in the proliferation and apoptosis of skin cells. The development of animal models has allowed a better understanding of these pathomechanisms, with the possibility of carrying out toxicological screening and drug development. In particular, the zebrafish (Danio rerio) has been established as one of the most important model organisms for cancer research...
July 18, 2017: International Journal of Molecular Sciences
https://www.readbyqxmd.com/read/28716076/klf8-regulates-left-right-asymmetric-patterning-through-modulation-of-kupffer-s-vesicle-morphogenesis-and-spaw-expression
#13
Che-Yi Lin, Ming-Yuan Tsai, Yu-Hsiu Liu, Yu-Fen Lu, Yi-Chung Chen, Yun-Ren Lai, Hsin-Chi Liao, Huang-Wei Lien, Chung-Hsiang Yang, Chang-Jen Huang, Sheng-Ping L Hwang
BACKGROUND: Although vertebrates are bilaterally symmetric organisms, their internal organs are distributed asymmetrically along a left-right axis. Disruption of left-right axis asymmetric patterning often occurs in human genetic disorders. In zebrafish embryos, Kupffer's vesicle, like the mouse node, breaks symmetry by inducing asymmetric expression of the Nodal-related gene, spaw, in the left lateral plate mesoderm (LPM). Spaw then stimulates transcription of itself and downstream genes, including lft1, lft2, and pitx2, specifically in the left side of the diencephalon, heart and LPM...
July 17, 2017: Journal of Biomedical Science
https://www.readbyqxmd.com/read/28715414/kinesin-1-promotes-chondrocyte-maintenance-during-skeletal-morphogenesis
#14
Adrian Santos-Ledo, Marina Garcia-Macia, Philip D Campbell, Marta Gronska, Florence L Marlow
During skeletal morphogenesis diverse mechanisms are used to support bone formation. This can be seen in the bones that require a cartilage template for their development. In mammals the cartilage template is removed, but in zebrafish the cartilage template persists and the bone mineralizes around the cartilage scaffold. Remodeling of unmineralized cartilage occurs via planar cell polarity (PCP) mediated cell rearrangements that contribute to lengthening of elements; however, the mechanisms that maintain the chondrocyte template that supports perichondral ossification remain unclear...
July 2017: PLoS Genetics
https://www.readbyqxmd.com/read/28714182/rare-coding-variants-in-mapk7-predispose-to-adolescent-idiopathic-scoliosis
#15
Wenjie Gao, Chong Chen, Taifeng Zhou, Shulan Yang, Bo Gao, Hang Zhou, Chengjie Lian, Zizhao Wu, Xianjian Qiu, Xiaoming Yang, Esam Alattar, Wentao Liu, Deying Su, Silong Sun, Yulan Chen, Kenneth M C Cheung, Youqiang Song, Keith K D Luk, Danny Chan, Pak Chung Sham, Chao Xing, Chiea Chuen Khor, Gabriel Liu, Junlin Yang, Yubin Deng, Dingjun Hao, Dongsheng Huang, Quan-Zhen Li, Caixia Xu, Peiqiang Su
Adolescent idiopathic scoliosis (AIS) is a complex genetic disorder characterized by three-dimensional spinal curvatures, affecting 2-3% of school age children, yet the causes underlying AIS are not well understood. Here, we firstly conducted a whole-exome sequencing and linkage analysis on a three-generation Chinese family with autosomal dominant (AD) AIS, then performed targeted sequencing in a discovery cohort comprising 20 AD-AIS families and 86 simplex patients, and finally identified three disease-associated missense variants (c...
July 17, 2017: Human Mutation
https://www.readbyqxmd.com/read/28713249/crispr-cas9-mediated-zebrafish-knock-in-as-a-novel-strategy-to-study-midbrain-hindbrain-boundary-development
#16
Gokul Kesavan, Avinash Chekuru, Anja Machate, Michael Brand
The midbrain-hindbrain boundary (MHB) acts as an organizer and controls the fate of neighboring cells to develop into either mesencephalic (midbrain) or metencephalic (hindbrain) cells by secreting signaling molecules like Wnt1 and Fgf8. The zebrafish is an excellent vertebrate model for studying MHB development due to the ease of gene manipulation and the possibility of following cellular dynamics and morphogenetic processes using live imaging. Currently, only very few reporter and/or Cre-driver lines are available to study gene expression at the MHB, hampering the understanding of MHB development, and traditional transgenic technologies using promoter/enhancer fragments or bacterial artificial chromosome (BAC)-mediated transgenesis often do not faithfully recapitulate endogenous expression patterns...
2017: Frontiers in Neuroanatomy
https://www.readbyqxmd.com/read/28701426/dynamics-of-in-vivo-asc-speck-formation
#17
Paola Kuri, Nicole L Schieber, Thomas Thumberger, Joachim Wittbrodt, Yannick Schwab, Maria Leptin
Activated danger or pathogen sensors trigger assembly of the inflammasome adaptor ASC into specks, large signaling platforms considered hallmarks of inflammasome activation. Because a lack of in vivo tools has prevented the study of endogenous ASC dynamics, we generated a live ASC reporter through CRISPR/Cas9 tagging of the endogenous gene in zebrafish. We see strong ASC expression in the skin and other epithelia that act as barriers to insult. A toxic stimulus triggered speck formation and rapid pyroptosis in keratinocytes in vivo...
July 12, 2017: Journal of Cell Biology
https://www.readbyqxmd.com/read/28692648/zebrafish-slc30a10-deficiency-revealed-a-novel-compensatory-mechanism-of-atp2c1-in-maintaining-manganese-homeostasis
#18
Zhidan Xia, Jiayu Wei, Yingniang Li, Jia Wang, Wenwen Li, Kai Wang, Xiaoli Hong, Lu Zhao, Caiyong Chen, Junxia Min, Fudi Wang
Recent studies found that mutations in the human SLC30A10 gene, which encodes a manganese (Mn) efflux transporter, are associated with hypermanganesemia with dystonia, polycythemia, and cirrhosis (HMDPC). However, the relationship between Mn metabolism and HMDPC is poorly understood, and no specific treatments are available for this disorder. Here, we generated two zebrafish slc30a10 mutant lines using the CRISPR/Cas9 system. Compared to wild-type animals, mutant adult animals developed significantly higher systemic Mn levels, and Mn accumulated in the brain and liver of mutant embryos in response to exogenous Mn...
July 2017: PLoS Genetics
https://www.readbyqxmd.com/read/28687732/the-cannabinoid-receptor-interacting-proteins-1-of-zebrafish-are-not-required-for-morphological-development-viability-or-fertility
#19
Laura Fin, Giorgia Bergamin, Roberto A Steiner, Simon M Hughes
The Cannabinoid Receptor Interacting Protein 1 (Cnrip1) was discovered as an interactor with the intracellular region of Cannabinoid Receptor 1 (CB1R, also known as Cnr1 or CB1). Functional assays in mouse show cannabinoid sensitivity changes and Cnrip1 has recently been suggested to control eye development in Xenopus laevis. Two Cnrip1 genes are described in zebrafish, cnrip1a and cnrip1b. In situ mRNA hybridisation revealed accumulation of mRNA encoding each gene primarily in brain and spinal cord, but also elsewhere...
July 7, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28678442/crispr-cas9-directed-gene-editing-for-the-generation-of-loss-of-function-mutants-in-high-throughput-zebrafish-f0-screens
#20
Sunita S Shankaran, Timothy J Dahlem, Brent W Bisgrove, H Joseph Yost, Martin Tristani-Firouzi
The ability to perform reverse genetics in the zebrafish model organism has been greatly advanced with the advent of the CRISPR (clustered regularly interspaced short palindromic repeats)/Cas9 (CRISPR-associated) system. The high level of efficiency in generating mutations when using the CRISPR/Cas9 system combined with the rapid generation time of the zebrafish model organism has made the possibility of performing F0 screens in this organism a reality. This unit describes a detailed protocol for performing an F0 screen using the CRISPR/Cas9 system in zebrafish starting with the design and production of custom CRISPR/Cas9 reagents for injection...
July 5, 2017: Current Protocols in Molecular Biology
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