Figure 1. Generation of Cacna1d^Ile772Met/+ mice.

(A) Topology of the Cacna1d gene (isoform ENSMUST00000238504.1) with Exons as blue boxes and intronic segments as thin lines. Introns longer than 2,000 bp are truncated. The sequence encoding the first 18 amino acids of exon 16 (encoding Ile772) is shown below. Substitution of ATG (Met; orange) for ATC (Ile; blue) was performed using CRISPR/Cas9 editing. The PAM site (gray background) as well as the gRNA (black box) used for mutagenesis are highlighted. (B) Sequencing of genomic DNA obtained from ear biopsies shows the heterozygous mutation. (C) Sequencing of cDNA from adrenal gland demonstrates expression of the mutant RNA.