ZFN-Scores

ZFN-Scores predicts and ranks zinc finger nuclease (ZFN) target sites and candidate ZFNs for genome engineering by scoring Extended Modular Assembly arrays using an ab initio B-score cutoff and empirical activity data.

Key Features:

• Extended Modular Assembly (MA): Uses the Extended MA approach for constructing ZFN arrays from modular one-finger components.
• One-finger archives: Utilizes publicly available one-finger archives as building blocks for array assembly.
• Array length evaluation: Evaluates three- to six-finger arrays to assess the effect of array length on ZFN activity.
• Drop-out linker scheme: Incorporates a drop-out linker scheme to enable rapid assessment of three- to six-finger combinations.
• Ab initio B-score cutoff: Applies an ab initio B-score cutoff of 15 to evaluate MA ZFNs.
• Empirical benchmarking: Analyzes 268 array variants to relate B-scores and array composition to observed activity.
• Reported performance metrics: Documents that six-finger MA arrays achieved activity at 71% of targeted loci in human and mouse cells and that roughly half of ZFNs exceeding the B-score cutoff were active.

Scientific Applications:

• Genome engineering: Selection and prioritization of ZFN target sites to induce double-strand breaks for targeted genome modification.
• ZFN design optimization: Informing construction of three- to six-finger ZFN arrays to improve nuclease activity.
• Mutation rate improvement: Guiding array selection to increase observed mutation rates in human and mouse cells.
• Comparative assembly strategies: Comparing modular assembly outcomes across array lengths and compositions.

Methodology:

Assembles candidate ZFN arrays using Extended Modular Assembly from one-finger archives, scores arrays with an ab initio B-score cutoff of 15, and analyzes 268 array variants including three- to six-finger combinations evaluated via a drop-out linker scheme.