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Genomic Scissors

ZINC FINGER NUCLEASES: Genomic Scissors ZFN EZEN FOK1 ZFN ZFN FOK1 DO000X 1. ZFN-mediated editing Engineered zinc finger nucleases (ZFN) are synthetic restriction enzyme that fuse to natural DNA-cleaving domains (Fok1). These dimers act as genomic scissors, cleaving specific sequences of DNA. FOK1 ZFN ZFN ZFN ZFN ZFN ZFN FOK1 2. Double-strand breaks Binding of ZFN-Fok1dimer to a specific DNA sequence cleaves both strands of DNA. E E E-E IIIIISTTAAT 9.99 Donor DNA Targeted gene addition Targeted mutagenesis • Non-homologous end-joining (NHEJ) • Homologous recombination (HR) NHEJ repairs double-strand breaks by joining the two ends together. This usually produces no mutation, but, when the repair is imperfect, base pairs can be inserted, deleted, or substituted, all of which result in mutagenesis. Introduction of a donor DNA in addition to ZFNS allows HR machinery to repair the double-strand break using the supplied DNA fragment as a template. This process permits targeted gene editing. Therapeutic Applications Scientists can make ZFNS that recognize virtually any gene target, allowing direct modulation of that gene's expression. ZFNS can disrupt, correct, or target the addition of a gene sequence, making this technology a powerful one for creating novel drug targets to treat disease. Because of their high specificity and selectivity, ZFNS may be needed in only low concentrations to provide its therapeutic benefits. This is opposite to small molecule and RNA-based approaches, which may have "off-target" effects and can be toxic at the concentrations that required to be therapeutically effective. ZENS can be used transiently to achieve its therapeutic effect because brief cellular expression of ZFNS causes permanent gene disruption, correction, or addition. RESOURCES: 1. Durai S, Mani M, Kandavelou K, Wu J, Porteus MH, Chandrasegaran S. Zinc finger nucleases: custom-designed molecular scissors for genome engineering of plant and mammalian cells. Nucleic Acids Res. 2005 Oct 26:33(18):5978-90. 2. Carroll D. Genome engineering with zinc-finger nucleases. Genetics. 2011 Aug:188(4):773-82. doi: 10.1534/genetics.111.131433. 3. Davis D, Stokoe D. Zinc finger nucleases as tools to understand and treat human diseases. BMC Med. 2010 Jul 1,8:42. doi: 10.1186/1741-7015-8-42. 4. © 2013 ZFN ZFN

Genomic Scissors

shared by J.L on Oct 25
Scientists can make ZFNs that recognize virtually any gene targets, allowing direct modulation of that gene's expression




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