Fig. 1

Replacement of the mouse Apoe gene with the human APOE gene in APOE-KI mice. a Genomic organization of the mouse Apoe gene containing exons 1–4. b The APOE-ε3 targeting construct containing the 5′ and 3′ arms of mouse homology interrupted by the human APOE-ε3 gene sequence. Exons 2 to 4 of the human sequence were flanked with loxP sites. Positive selection markers were flanked by FRT (Neomycin resistance – NeoR) and F3 (Puromycin resistance – PuroR) sites and inserted downstream of the proximal loxP site and upstream of the distal loxP site, respectively. c Homologous recombinant clones were isolated using double positive (NeoR and PuroR) and negative (Thymidine kinase - TK) selections. d The constitutive humanized/conditional knock-out alleles were achieved after in vivo Flp-mediated removal of the selection markers. The newly introduced human APOE gene is expressed under control of the endogenous Apoe promoter. e Constitutive knock-out allele is achieved when the loxP-flanked region is removed by Cre-recombinase. f PBS-soluble apoE protein concentration was measured in cortical brain homogenates from APOE-KI mice at 21 days (p = 0.0004, F = 15.77). g PBS-soluble apoE protein concentration was measured in cortical brain homogenates from APOE-KI mice at 3 months of age (p = 0.0043, F = 8.880). h Guanidine-soluble apoE protein concentration was also measured in cortical brain homogenates from APOE-KI mice at 3 months of age (p = 0.0144, F = 6.169). i The same brain homogenates were subjected to Western blot analysis for apoE using antibody HJ15.7. White arrowhead = sialylated apoE (MW ~ 35.3 kDa). Black arrow = non-sialylated apoE (MW ~ 33.6 kDa). N = 2 males, 2 females per genotype. *p < 0.05, ***p < 0.001. A one-way ANOVA was used to assess significance between more than two groups, and Bonferroni’s post-hoc test was used to test for differences between each of the groups. All values are reported as mean ± SEM. N = 5 per genotype for f, g, and h, with approximately equal numbers of males and females