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ZFNGenome: A Comprehensive Resource for Locating Zinc Finger Nuclease Target Sites in Model Organisms


ZFNGenome: A Comprehensive Resource for Locating Zinc Finger Nuclease Target Sites in Model Organisms

Deepak Reyon et al. BMC Genomics.


Background: Zinc 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.

Description: ZFNGenome 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.

Conclusions: 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.ZFNGenome is freely available at: or at the Zinc Finger Consortium website:


Figure 1
Figure 1
ZFNs generate site-specific double-stranded breaks that can be used for homologous recombination or mutagenesis. (A) ZFNs are composed of two arrays that recognize 9-12 base pairs each. Two arrays with three fingers, F1-F2-F3, that recognize nine base pairs each are shown. Each array is fused to one half of a nonspecific FokI endonuclease (green). Upon dimerization, the FokI endonuclease is activated and creates a double-stranded break at sites flanked by the DNA binding sites recognized by the zinc finger arrays. Scissors and arrows denote the cut sites. (B) In most cells, double-stranded breaks (DSBs) are repaired by one of two major pathways. If a donor template is available, homologous recombination can result in engineered nucleotide substitutions at the target site (left). Alternatively, DSBs can be repaired by non-homologous end-joining, an error-prone mechanism that frequently results in small deletions or insertions at the site of the DSB (right).
Figure 2
Figure 2
An overview of the ZFNGenome architecture. ZFNGenome creates a user-friendly interface for visualizing all potential ZFN target sites in seven model organisms by integrating the genomic information from genome browsers of sequenced and annotated genomes with the tools of the Zinc Finger Consortium. This interface allows researchers using these model organisms to easily determine whether ZFNs are available for the design and execution of targeted genome modifications.
Figure 3
Figure 3
Examples of resources available in ZFNGenome. (A) The ZFNGenome Homepage is shown. From here, the user can select a model organism from the seven shown in the left hand column. In addition, links to the ZFNGenome Tutorial and Help pages are provided. (B) A screenshot of the result of a search of the S. cerevisiae gene YLR219W is displayed. Key areas of the browser include the search box and the "Scroll/Zoom" areas at the top. The "Overview" and "Detail" panels serve as controls for visualizing the genome. This search shows the single coding region of this gene has 17 potential ZFN target sites, color-coding according to their "uniqueness" and "ZiFOpT" scores (see text). Additional information on each of the tracks can be obtained by clicking on details of the track. For example, clicking on one of the OPEN Zinc Finger Nuclease Sites links the user to details about that specific ZFN target. (C) The ZFNGenome Tutorial offers instructions on navigating the database. The Tutorial can be accessed from the Homepage or from any GBrowse page within ZFNGenome. Help and Instruction links are provided from the GBrowse pages. (D) Clicking on a ZFN target site opens a new window that provides links to ZiFDB, which provides additional information for each zinc finger array, ZiFiT the zinc finger design software that includes the OPEN design method and zinc finger pools, and the BLAST server at National Center for Biotechnology Information (NCBI).

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