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. 2011 Oct 14;21(2):384–393. doi: 10.1093/hmg/ddr473

Figure 1.

Figure 1.

Generation of Col8a2 Q455K knock-in mice. (A) Map of targeting strategy. The targeting vector containing a portion of Col8a2 intron 2, the entire exon 3 (Ex3) and intron 3 and a portion of exon 4 on mouse chromosome (chr) 4 was produced as described in the Materials and Methods. The targeting vector contained the Col8a2 Q455K mutation (asterisk) in exon 4 and an additional diagnostic MfeI site in intron 3. Chimeric animals were bred to heterozygosity and subsequently crossed with a cre-recombinase expressing line. (B) Southern analysis of MfeI-digested genomic DNA from two different (i) F1 WT, (ii) F1 Col8a2WT/Q455K heterozygous and (iii) F2 Col8a2WT/Q455K heterozygous animals. F1 WT mice show an 18.1 kb band (i) and F1 heterozygotes show an additional 6.1 kb band for the mutant allele (ii). F2 heterozygotes show cre-mediated excision of the pGK-neomycin cassette and a smaller 4.2 kb mutant band (iii). (C) Sequence analysis of Col8a2WT/Q455K mutant mice. Both the WT and mutant sequences (C/A) are seen with equal chromatogram signal intensity (arrow). (D) Fnu4HI restriction enzyme digestion of PCR fragments showing two different Col8a2Q455K/Q455K (MUT, 1–2) and WT (3–4) mice. The Q455K mutation results in a 574bp band, and the WT sequence results in a 384 bp band.