Figure 1.
The OPEN Zinc Finger Selection Method.
(A) Schematic overview of OPEN selection for a target DNA site. Zinc finger domains are shown as spheres and associated 3 bp subsites as rectangles. Details provided in the text and in Maeder et al., Mol. Cell 2008. (B) Schematic of the bacterial two-hybrid (B2H) system. ZFP = zinc-finger protein. X and Y = arbitrary interacting proteins.
Table 1.
Recognition helix (RH) amino acid sequences and B2H activities of zinc finger arrays for endogenous zebrafish gene targets.
Figure 2.
Toxicity and teratogenicity of OPEN and B1H-selected ZFNs in zebrafish embryos.
Percentages of dead, deformed (“monster”), and normal embryos following injection with the amounts of ZFN RNAs indicated are shown. Percentages were calculated from the number of embryos (n) indicated.
Figure 3.
Frequencies and sequences of ZFN-induced mutations in somatic zebrafish cells.
For each gene targeted by ZFNs, the wild-type sequence is shown at the top with ZFN binding sites marked. Deletions are indicated by grey highlighted red dashes and insertions by blue highlighted lower case blue letters. The number of times each wild-type mutant allele was isolated is shown in brackets.
Figure 4.
Sequences of ZFN-induced mutations transmitted through the germline.
For each target gene, the wild-type sequence is shown at the top with ZFN binding sites marked and the mutated alleles found in founder progeny are shown below the wild-type sequence. Each mutant sequence shown was isolated from progeny of different founders. Deletions are indicated by grey highlighted red dashes and insertions by blue highlighted lower case blue letters.
Table 2.
Frequencies of mutations from founder analysis
Table 3.
Summary of potential OPEN ZFN target sites identified in zebrafish transcripts