Mecp2^em1Bird

A targeting vector, an sgRNA and CRISPR/Cas9 technology were used to modify the locus as follows: a loxP site, a neomycin resistance gene cassette, a transcriptional STOP cassette and a second loxP site, all inserted into intron 2, a modified exon 3 where the sequence for codons 30-87 (splice variant e1) or 13-70 (splice variant e2) was deleted, sequence for linker peptide CKDPPVAT, and the EGFP gene. The neo-STOP cassette was subsequently removed through cre-mediated recombination. This leaves a knockin allele where the endogenous gene is replaced with a truncated version (with sequences immediately upstream of the methyl-CpG binding domain (MBD) deleted) fused to a fluorescent marker. The truncated protein retains 88% of the wild-type sequence. (J:252845)