Home LiteratureArticle Details
PMID: 18657511 Published · ppublish English Journal Article Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, Non-P.H.S.

Rapid "open-source" engineering of customized zinc-finger nucleases for highly efficient gene modification.

Molecular cell ·Vol. 31 ·No. 2 ·2008-07-25 ·Pages 294-301

Maeder ML, Thibodeau-Beganny S, Osiak A, Wright DA, Anthony RM, Eichtinger M, Jiang T, Foley JE, Winfrey RJ, Townsend JA, Unger-Wallace E, Sander JD, Müller-Lerch F, Fu F, Pearlberg J, Göbel C, Dassie JP, Pruett-Miller SM, Porteus MH, Sgroi DC, Iafrate AJ, Dobbs D, McCray PB, Cathomen T, Voytas DF, Joung JK

Abstract

Custom-made zinc-finger nucleases (ZFNs) can induce targeted genome modifications with high efficiency in cell types including Drosophila, C. elegans, plants, and humans. A bottleneck in the application of ZFN technology has been the generation of highly specific engineered zinc-finger arrays. Here we describe OPEN (Oligomerized Pool ENgineering), a rapid, publicly available strategy for constructing multifinger arrays, which we show is more effective than the previously published modular assembly method. We used OPEN to construct 37 highly active ZFN pairs which induced targeted alterations with high efficiencies (1%-50%) at 11 different target sites located within three endogenous human genes (VEGF-A, HoxB13, and CFTR), an endogenous plant gene (tobacco SuRA), and a chromosomally integrated EGFP reporter gene. In summary, OPEN provides an "open-source" method for rapidly engineering highly active zinc-finger arrays, thereby enabling broader practice, development, and application of ZFN technology for biological research and gene therapy.

MeSH Terms
Base Sequence Endonucleases/metabolism,toxicity Gene Targeting Genetic Engineering/methods Green Fluorescent Proteins/genetics Humans K562 Cells Molecular Sequence Data Mutagenesis Mutation/genetics Protein Conformation Zinc Fingers
Chemicals
enhanced green fluorescent protein Green Fluorescent Proteins Endonucleases
Authors & Affiliations
26 authors, click to expand affiliations / ORCID
Maeder Morgan L
Molecular Pathology Unit and Center for Cancer Research, Massachusetts General Hospital, Charlestown, MA 02129, USA.
Thibodeau-Beganny Stacey
Osiak Anna
Wright David A
Anthony Reshma M
Eichtinger Magdalena
Jiang Tao
Foley Jonathan E
Winfrey Ronnie J
Townsend Jeffrey A
Unger-Wallace Erica
Sander Jeffry D
Müller-Lerch Felix
Fu Fengli
Pearlberg Joseph
Göbel Carl
Dassie Justin P
Pruett-Miller Shondra M
Porteus Matthew H
Sgroi Dennis C
Iafrate A John
Dobbs Drena
McCray Paul B
Cathomen Toni
Voytas Daniel F
Joung J Keith
References (38)
38 references, click to expand
  1. Highly efficient endogenous human gene correction using designed zinc-finger nucleases.
    Nature. 2005 Jun 2;435(7042):646-51 PMID: 15806097
  2. Engineering Cys2His2 zinc finger domains using a bacterial cell-based two-hybrid selection system.
    Methods Mol Biol. 2007;408:317-34 PMID: 18314590
  3. Gene targeting using zinc finger nucleases.
    Nat Biotechnol. 2005 Aug;23(8):967-73 PMID: 16082368
  4. An improved zinc-finger nuclease architecture for highly specific genome editing.
    Nat Biotechnol. 2007 Jul;25(7):778-85 PMID: 17603475
  5. Engineering polydactyl zinc-finger transcription factors.
    Nat Biotechnol. 2002 Feb;20(2):135-41 PMID: 11821858
  6. Induction and repair of zinc-finger nuclease-targeted double-strand breaks in Caenorhabditis elegans somatic cells.
    Proc Natl Acad Sci U S A. 2006 Oct 31;103(44):16370-5 PMID: 17060623
  7. Gene editing in human stem cells using zinc finger nucleases and integrase-defective lentiviral vector delivery.
    Nat Biotechnol. 2007 Nov;25(11):1298-306 PMID: 17965707
  8. Targeted mutagenesis using zinc-finger nucleases in Arabidopsis.
    Proc Natl Acad Sci U S A. 2005 Feb 8;102(6):2232-7 PMID: 15677315
  9. Enhancing gene targeting with designed zinc finger nucleases.
    Science. 2003 May 2;300(5620):764 PMID: 12730594
  10. Analysis of zinc fingers optimized via phage display: evaluating the utility of a recognition code.
    J Mol Biol. 1999 Feb 5;285(5):1917-34 PMID: 9925775
  11. Targeted chromosomal cleavage and mutagenesis in Drosophila using zinc-finger nucleases.
    Genetics. 2002 Jul;161(3):1169-75 PMID: 12136019
  12. Unexpected failure rates for modular assembly of engineered zinc fingers.
    Nat Methods. 2008 May;5(5):374-5 PMID: 18446154
  13. Genetic manipulation of genomes with rare-cutting endonucleases.
    Trends Genet. 1996 Jun;12(6):224-8 PMID: 8928227
  14. A general strategy for selecting high-affinity zinc finger proteins for diverse DNA target sites.
    Science. 1997 Jan 31;275(5300):657-61 PMID: 9005850
  15. Structure-based redesign of the dimerization interface reduces the toxicity of zinc-finger nucleases.
    Nat Biotechnol. 2007 Jul;25(7):786-93 PMID: 17603476
  16. A bacterial two-hybrid selection system for studying protein-DNA and protein-protein interactions.
    Proc Natl Acad Sci U S A. 2000 Jun 20;97(13):7382-7 PMID: 10852947
  17. Highly specific zinc finger proteins obtained by directed domain shuffling and cell-based selection.
    Proc Natl Acad Sci U S A. 2003 Oct 14;100(21):12271-6 PMID: 14527993
  18. Evaluation of a modular strategy for the construction of novel polydactyl zinc finger DNA-binding proteins.
    Biochemistry. 2003 Feb 25;42(7):2137-48 PMID: 12590603
  19. Efficient gene targeting in Drosophila with zinc-finger nucleases.
    Genetics. 2006 Apr;172(4):2391-403 PMID: 16452139
  20. Zinc Finger Tools: custom DNA-binding domains for transcription factors and nucleases.
    Nucleic Acids Res. 2006 Jul 1;34(Web Server issue):W516-23 PMID: 16845061
  21. Synergy between adjacent zinc fingers in sequence-specific DNA recognition.
    Proc Natl Acad Sci U S A. 1997 May 27;94(11):5617-21 PMID: 9159121
  22. Zif268 protein-DNA complex refined at 1.6 A: a model system for understanding zinc finger-DNA interactions.
    Structure. 1996 Oct 15;4(10):1171-80 PMID: 8939742
  23. Zinc Finger Targeter (ZiFiT): an engineered zinc finger/target site design tool.
    Nucleic Acids Res. 2007 Jul;35(Web Server issue):W599-605 PMID: 17526515
  24. Zinc-finger nucleases: the next generation emerges.
    Mol Ther. 2008 Jul;16(7):1200-7 PMID: 18545224
  25. A rapid, generally applicable method to engineer zinc fingers illustrated by targeting the HIV-1 promoter.
    Nat Biotechnol. 2001 Jul;19(7):656-60 PMID: 11433278
  26. Validated zinc finger protein designs for all 16 GNN DNA triplet targets.
    J Biol Chem. 2002 Feb 8;277(6):3850-6 PMID: 11726671
  27. DNA-binding specificity is a major determinant of the activity and toxicity of zinc-finger nucleases.
    Mol Ther. 2008 Feb;16(2):352-8 PMID: 18026168
  28. Human zinc fingers as building blocks in the construction of artificial transcription factors.
    Nat Biotechnol. 2003 Mar;21(3):275-80 PMID: 12592413
  29. Beyond the "recognition code": structures of two Cys2His2 zinc finger/TATA box complexes.
    Structure. 2001 Aug;9(8):717-23 PMID: 11587646
  30. Stimulation of homologous recombination through targeted cleavage by chimeric nucleases.
    Mol Cell Biol. 2001 Jan;21(1):289-97 PMID: 11113203
  31. Targeted gene knockout in mammalian cells by using engineered zinc-finger nucleases.
    Proc Natl Acad Sci U S A. 2008 Apr 15;105(15):5809-14 PMID: 18359850
  32. Chimeric nucleases stimulate gene targeting in human cells.
    Science. 2003 May 2;300(5620):763 PMID: 12730593
  33. Targeted gene addition into a specified location in the human genome using designed zinc finger nucleases.
    Proc Natl Acad Sci U S A. 2007 Feb 27;104(9):3055-60 PMID: 17360608
  34. Custom zinc-finger nucleases for use in human cells.
    Mol Ther. 2005 Oct;12(4):610-7 PMID: 16039907
  35. 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 PMID: 16251401
  36. Comparison of zinc finger nucleases for use in gene targeting in mammalian cells.
    Mol Ther. 2008 Apr;16(4):707-17 PMID: 18334988
  37. High-frequency homologous recombination in plants mediated by zinc-finger nucleases.
    Plant J. 2005 Nov;44(4):693-705 PMID: 16262717
  38. AAV vectors for gene correction.
    Curr Opin Mol Ther. 2004 Aug;6(4):360-6 PMID: 15468594
Article Info
Journal
Molecular cell
Abbr.
Mol Cell
ISSN
1097-4164
DOI
10.1016/j.molcel.2008.06.016
Published
2008-07-25
Pages
294-301
Language
English
Region
United States
NLM ID
9802571
PMCID
PMC2535758
Subset
IM
Grants
NIGMS NIH HHS · R01 GM069906-03 · United States
NIGMS NIH HHS · R01GM069906 · United States
NCI NIH HHS · R01CA112021 · United States
NIGMS NIH HHS · R24 GM078369-02 · United States
NIGMS NIH HHS · R01 GM069906 · United States
NIGMS NIH HHS · R24 GM078369 · United States
NCI NIH HHS · R01 CA112021 · United States
NCRR NIH HHS · R21 RR024189-01 · United States
NIH HHS · DP1 OD006862 · United States
NIGMS NIH HHS · R01 GM069906-05 · United States
NIGMS NIH HHS · R24GM078369 · United States
NIGMS NIH HHS · R24 GM078369-01A1 · United States
NCRR NIH HHS · R21RR024189 · United States
NCRR NIH HHS · R21 RR024189 · United States
NHLBI NIH HHS · R01 HL079295 · United States
NIGMS NIH HHS · R01 GM069906-04 · United States
External Links
PubMed source DOI: 10.1016/j.molcel.2008.06.016 PMC Full Text View journal details
Analysis Services
Analysis Services

Contact

No. 2 Wenbo Road, Zhangqiu District, Jinan, Shandong

Qilu Normal University · Genelibs Bioinformatics Lab

750 Shunhua Rd, Jinan

2F, Bldg F, University Science Park

Tel: 0531-88819269

WeChat Official Account

Follow our WeChat subscription account for real-time updates and the latest in medical and biological research.


Business Email

E-mail: [email protected]

Copyright © 2026.Genelibs Jinan Honghui Biotechnology Co., Ltd. 鲁ICP备20007159号-2