Figure 1
From: Fat mass- and obesity-associated gene Fto affects the dietary response in mouse white adipose tissue
Disruption of the mouse Fto gene.
WT wild type; Fto-KO Fto-knockout. (a) An embryonic stem cell line containing a ‘knock-in-first’ construct targeting the second intron of mouse Fto gene, disrupting the endogenous transcription and leading to production of truncated mRNA, was used to generate Fto-KO mouse line. Arrows represent the primer binding sites used in the genotyping; E1–E9 exons 1–9; En2SA En2 splice acceptor; β-geo fusion of β galactosidase and neomycin resistance genes; pA polyadenylation signal; UTR untranslated region; Frt flippase recognition target site; loxP locus of crossover in P1 site. (b) Genotyping of WT and Fto-KO mice by PCR. A 500-bp fragment was produced from the WT allele with Fto-specific primers which lack the Fto-KO allele due to the construct. Primers targeting the neomycin resistance gene (Neo) produced a 475-bp fragment from the Fto-KO construct which was not present in the WT allele. (c) Absence of FTO protein in Fto-KO mice was confirmed with Western blot by using an antibody against mouse FTO and donkey anti-rat IgG as a secondary antibody. Recombinant FTO (Rec. Fto) was used as a positive control (60 kDa) which was run in the same gel and under same experimental conditions as the samples. Dotted lines indicate the cropping lines.
