Figure 3. mNG2₁₁ tagging by CRISPR/Cas-directed gene editing.

A. Schematic of CRISPR/Cas-directed mNG2₁₁ insertion into target genes. Purple, endogenous exon sequence. Yellow, linker (LK). Green, mNG2₁₁. ATG, start codon. Arrows denote primers used in B. B. mNG2₁₁ insertion was assessed by PCR. The primers used correspond to the arrows shown in A. C. Amino acid sequences of wild-type, predicted mNG2₁₁ fusions, and recovered alleles for Tubb4b and Krt8. Mismatches between the predicted and recovered sequences are highlighted in red. Asterisks, stop codons. D–I. Representative images of mNG2₁₁-tubb4b (D-F) and krt8-mNG2₁₁ (G–I) embryos injected with mNG2_1–10 mRNA (D–E, G–H) or uninjected (F, I). Images were acquired at 24 hours post-fertilization. Images in F and I have been overexposed to emphasize lack of fluorescence. Scale bars, 50 μm.