Abstract

BackgroundZinc Finger Nucleases (ZFNs) have tremendous potential as tools to facilitate genomic modifications, such as precise gene knockouts or gene replacements by homologous recombination. ZFNs can be used to advance both basic research and clinical applications, including gene therapy. Recently, the ability to engineer ZFNs that target any desired genomic DNA sequence with high fidelity has improved significantly with the introduction of rapid, robust, and publicly available techniques for ZFN design such as the Oligomerized Pool ENgineering (OPEN) method. The motivation for this study is to make resources for genome modifications using OPEN-generated ZFNs more accessible to researchers by creating a user-friendly interface that identifies and provides quality scores for all potential ZFN target sites in the complete genomes of several model organisms.DescriptionZFNGenome is a GBrowse-based tool for identifying and visualizing potential target sites for OPEN-generated ZFNs. ZFNGenome currently includes a total of more than 11.6 million potential ZFN target sites, mapped within the fully sequenced genomes of seven model organisms; S. cerevisiae, C. reinhardtii, A. thaliana, D. melanogaster, D. rerio, C. elegans, and H. sapiens and can be visualized within the flexible GBrowse environment. Additional model organisms will be included in future updates. ZFNGenome provides information about each potential ZFN target site, including its chromosomal location and position relative to transcription initiation site(s). Users can query ZFNGenome using several different criteria (e.g., gene ID, transcript ID, target site sequence). Tracks in ZFNGenome also provide "uniqueness" and ZiFOpT (Zinc Finger OPEN Targeter) "confidence" scores that estimate the likelihood that a chosen ZFN target site will function in vivo. ZFNGenome is dynamically linked to ZiFDB, allowing users access to all available information about zinc finger reagents, such as the effectiveness of a given ZFN in creating double-stranded breaks.ConclusionsZFNGenome provides a user-friendly interface that allows researchers to access resources and information regarding genomic target sites for engineered ZFNs in seven model organisms. This genome-wide database of potential ZFN target sites should greatly facilitate the utilization of ZFNs in both basic and clinical research.ZFNGenome is freely available at: http://bindr.gdcb.iastate.edu/ZFNGenome or at the Zinc Finger Consortium website: http://www.zincfingers.org/.

Highlights

  • Zinc Finger Nucleases (ZFNs) have tremendous potential as tools to facilitate genomic modifications, such as precise gene knockouts or gene replacements by homologous recombination

  • ZFNGenome provides a user-friendly interface that allows researchers to access resources and information regarding genomic target sites for engineered ZFNs in seven model organisms. This genome-wide database of potential ZFN target sites should greatly facilitate the utilization of ZFNs in both basic and clinical research

  • Engineered ZFNs consist of two zinc finger arrays (ZFAs), each of which is fused to a single subunit of a non-specific endonuclease, such as the nuclease domain from the FokI enzyme, which becomes active upon dimerization [20,21]

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Summary

Conclusions

OPEN is a robust, publicly available, experimental platform for the generation of engineered ZFNs that function with high specificity in vivo. ZFNGenome was developed to enhance and broaden the applicability of ZFNs for genomic modification by providing an online resource that contains all potential target sites for OPEN-generated ZFNs in the sequenced genomes of several model organisms. ZFNGenome has a userfriendly interface and is seamlessly integrated with other publicly available Zinc Finger Consortium resources, such as ZiFiT, ZiFDB, and ZiFOpT. List of Abbreviations Used OPEN: Oligomerized Pool ENgineering; ZF: Zinc Finger; ZFA: Zinc Finger Array; ZFP: Zinc Finger Protein; ZFN: Zinc Finger Nuclease; ZiFOpT: Zinc Finger OPEN Targeter; DSB: double stranded breaks; HR: homologous recombination; NHEJ: non-homologous end joining; GMOD: generic model organism database project; SVMs: support vector machines; ZiFDB: Zinc Finger Database

Background
Utility and Discussion
Findings
Carroll D
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