Figure 2. Generation of mice harboring the Snap23 ^fl or Snap23 ^Δ allele.

(A and B) Southern blot analysis of genomic DNA from targeted Snap23 ^Neo+fl/wt ES cell clones. (A) EcoRI-digested genomic DNA was hybridized with a 5′ probe as shown in Figure 1A. Southern blot analysis of EcoRI-digested genomic DNA using the 5′ probe revealed a 3.4 kb or 2.5 kb fragment in the Snap23 ^wt or Snap23 ^Neo+fl targeted allele, respectively. (B) Southern blot analysis of ClaI/KpnI-digested genomic DNA using a 3′ probe revealed a 9.6 kb or 6 kb fragment from the Snap23 ^wt or Snap23 ^Neo+fl targeted allele, respectively. (C) Genomic PCR from Snap23 ^Neo+fl/wt heterozygous mice using PCR Primer set 1. The expected size of PCR products is 266 bp from the Snap23 ^wt allele and 400 bp for the Snap23 ^Neo+fl allele. (D) Genotyping was performed using PCR Primer set 2 to identify Snap23 mosaic mice. Three PCR products of 266 bp, 400 bp, and 492 bp were obtained from mice with ear tag numbers 173312 or 173313, indicating these mice harbor a mixed mosaic genotype (F2) depicted in Figure 1A. Mice with ear tag number 173314 or 173315 yielded a single PCR fragment of 266 bp, indicating these possess either the Snap23 ^wt or Snap23 ^Neo+Δ allele. The mosaic EIIa-Cre⁺ male mouse 173312 was mated with a female C57BL/6 mouse and one of the pups (F3; ear tag number 182234) was found to be EIIa-Cre^- and possessed the Snap23 ^fl allele. Genomic PCR from this mouse revealed only two PCR fragments corresponding the Snap23 ^wt and Snap23^fl allele, indicating that this was a Snap23 floxed exon 2 heterozygous mouse.