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  • The Zinc Finger Consortium

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  • The Zinc Finger Consortium
    ZFNs are an extremely powerful tool for performing targeted genomic manipulation in a variety of cell types from Drosophila to humans ZFNs consist of an engineered DNA binding zinc finger domain linked to a non specific endonuclease domain and can introduce double stranded breaks DSBs that stimulate both homologous and non homologous recombination processes that can be harnessed to perform genomic manipulation The capability to alter any genomic locus of

    Original URL path: http://www.zincfingers.org/default2.htm (2016-02-10)
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  • The Zinc Finger Consortium | Scientific Background
    be enormously useful for biological research and molecular therapeutics Highly efficient targeted genome manipulation induced by ZFNs Gene targeting is a method to repair or inactivate any desired gene of interest Gene targeting strategies use the introduction of a double stranded break DSB into a genomic locus to enhance the efficiency of recombination with an exogenously introduced homologous DNA repair template Figure 1 DSBs can stimulate recombination efficiency several thousand fold approaching gene targeting frequencies as high as 50 ZFNs can be used to introduce targeted DSBs ZFNs consist of a DNA binding zinc finger domain composed of three to six fingers covalently linked to the non specific DNA cleavage domain of the bacterial FokI restriction endonuclease Figure 2 left panel ZFNs can bind as dimers to their target DNA sites with each monomer using its zinc finger domain to recognize a half site Figure 2 right panel Dimerization of ZFNs is mediated by the FokI cleavage domain which cleaves within a five or six base pair spacer sequence that separates the two inverted half sites Figure 2 right panel Importantly because the DNA binding specificities of zinc finger domains can in principle be re engineered using one of various

    Original URL path: http://www.zincfingers.org/scientific-background.htm (2016-02-10)
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  • The Zinc Finger Consortium | Consortium Members
    of lymphohematopoietic cell transfer Blazar lab webpage Linzhao Cheng Ph D Johns Hopkins University School of Medicine Baltimore MD Expertise in genetic manipulation of human stem cells Cheng lab webpage George M Church Ph D Harvard Medical School Boston MA Expertise in high throughput sequencing homologous recombination synthetic biology and personal genomics Church lab webpage Drena L Dobbs Ph D Iowa State University Ames IA Expertise in bioinformatics and computational biology Dobbs lab webpage Mark Isalan Ph D EMBL CRG Systems Biology Programme Barcelona Spain Expertise in the engineering selection and design of zinc finger domains Isalan lab webpage Paul B McCray Jr M D University of Iowa Iowa City IA Expertise in cystic fibrosis and gene transfer to respiratory epithelia McCray lab webpage Randall Peterson Ph D Massachusetts General Hospital and Harvard Medical School Expertise in zebrafish technologies and chemical biology Peterson lab webpage Matthew Porteus M D Ph D University of Texas Southwestern Medical Center Dallas TX Expertise in the use of ZFNs for gene targeting in mammalian cells Porteus lab webpage Jakub Tolar M D Ph D University of Minnesota Minneapolis MN Expertise clinically relevant stem cell gene correction of congenital disorders Tolar web page Éva Scheuring

    Original URL path: http://www.zincfingers.org/consortium-members.htm (2016-02-10)
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  • The Zinc Finger Consortium | Publications
    specificity from polyzinc finger peptides by using strings of two finger units Proc Natl Acad Sci U S A 98 1437 41 Pabo C O Peisach E Grant R A 2001 Design and selection of novel Cys2His2 zinc finger proteins Annu Rev Biochem 70 313 40 Pavletich N P Pabo C O 1991 Zinc finger DNA recognition crystal structure of a Zif268 DNA complex at 2 1 A Science 252 809 17 Rebar E J Greisman H A Pabo C O 1996 Phage display methods for selecting zinc finger proteins with novel DNA binding specificities Methods Enzymol 267 129 49 Rebar E J Pabo C O 1994 Zinc finger phage affinity selection of fingers with new DNA binding specificities Science 263 671 3 Segal D J Crotty J W Bhakta M S Barbas C F 3rd Horton N C 2006 Structure of Aart a designed six finger zinc finger peptide bound to DNA J Mol Biol 363 405 421 Segal D J Dreier B Beerli R R Barbas C F 3rd 1999 Toward controlling gene expression at will selection and design of zinc finger domains recognizing each of the 5 GNN 3 DNA target sequences Proc Natl Acad Sci U S A 96 2758 63 Thibodeau Beganny S and Joung J K 2007 Engineering Cys2His2 zinc finger domains using a bacterial cell based two hybrid selection system Methods Mol Biol 408 317 334 Wolfe S A Grant R A Elrod Erickson M Pabo C O 2001 Beyond the recognition code structures of two Cys2His2 zinc finger TATA box complexes Structure Camb 9 717 23 Wolfe S A Greisman H A Ramm E I Pabo C O 1999 Analysis of zinc fingers optimized via phage display evaluating the utility of a recognition code J Mol Biol 285 1917 34 Wolfe S A Nekludova L Pabo C O 2000 DNA recognition by Cys2His2 zinc finger proteins Annu Rev Biophys Biomol Struct 29 183 212 Wright D A Thibodeau Beganny S Sander J D Winfrey R J Hirsh A S Eichtinger M Fu F Porteus M H Dobbs D Voytas D F Joung J K 2006 Standardized reagents and protocols for engineering zinc finger nucleases by modular assembly Nature Protocols 1 1637 1652 Wu H Yang W P Barbas C F 3rd 1995 Building zinc fingers by selection toward a therapeutic application Proc Natl Acad Sci U S A 92 344 8 Zinc Finger Nucleases Alwin S Gere M B Guhl E Effertz K Barbas C F 3rd Segal D J Weitzman M D Cathomen T 2005 Custom Zinc Finger Nucleases for Use in Human Cells Mol Ther 12 610 617 Beumer K Bhattacharyya G Bibikova M Trautman J K Carroll D 2006 Efficient gene targeting in Drosophila with zinc finger nucleases Genetics 172 2391 2403 Bibikova M Beumer K Trautman J K Carroll D 2003 Enhancing gene targeting with designed zinc finger nucleases Science 300 764 Bibikova M Carroll D Segal D J Trautman J K Smith J Kim Y G Chandrasegaran

    Original URL path: http://www.zincfingers.org/publications.htm (2016-02-10)
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  • The Zinc Finger Consortium | Software Tools
    ZiFiT web accessible software for the design of engineered zinc finger arrays ZiFDB web accessible database of zinc fingers and engineered zinc finger arrays ZFNGenome genome scale resource for identifying

    Original URL path: http://www.zincfingers.org/software-tools.htm (2016-02-10)
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  • The Zinc Finger Consortium | Protocols
    and Resources Context Dependent Assembly CoDA method for engineering zinc finger arrays Oligomerized Pool Engineering OPEN method for engineering zinc finger arrays Modular Assembly method for engineering zinc finger arrays ZFN mediated mutagenesis of endogenous zebrafish genes High throughput beta

    Original URL path: http://www.zincfingers.org/protocols.htm (2016-02-10)
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  • The Zinc Finger Consortium | Reagents
    Visit the Joung Lab Page Lab Information and Resources Oligomerized Pool Engineering OPEN reagents Modular assembly zinc finger engineering reagents Zinc Finger Nuclease ZFN expression vectors

    Original URL path: http://www.zincfingers.org/reagents.htm (2016-02-10)
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