Basic Information
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Targeting strategy
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Gene targeting strategy for B-hLAT1 mice.
Exons 1-10 of the mouse Slc7a5 gene (encoding the full-length protein) were replaced by human SLC7A5 exons 1-10 to generate humanized B-hLAT1 mice.
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mRNA expression analysis
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Strain specific analysis of LAT1 gene expression in wild type (WT) mice and B-hLAT1 mice by RT-PCR. Mouse Slc7a5 mRNA was detectable only in brain of WT mice (+/+). Human SLC7A5 mRNA was detectable in homozygous B-hLAT1 mice (H/H) but not in WT mice (+/+).
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Protein expression analysis
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Strain specific LAT1 expression analysis in homozygous B-hLAT1 mice by western blot. Brain was collected from wild type (WT) mice (+/+) and homozygous B-hLAT1 mice (H/H), and analyzed by western blot with anti-LAT1 antibody. LAT1 was detectable in WT mice (+/+) and homozygous B-hLAT1 mice (H/H) due to the cross-reactivity of antibodies. Human LAT1 was exclusively detectable in homozygous B-hLAT1 mice (H/H).
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Protein expression analysis in brain
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Strain specific LAT1 expression analysis in homozygous B-hLAT1 mice by Immunofluorescence staining.
Brain was collected from wild type (WT) mice and homozygous B-hLAT1 mice (female,8 week old) and processed into paraffin sections. The paraffin sections were stained using an species-specific anti-human LAT1 antibody (green). Brain tissues were co-stained with an anti-mouse CD31 antibody to visualize microvascular endothelial cells (red). Representative results of WT mice (A) and homozygous B-hLAT1 mice (B) are shown. The results indicated that human LAT1 was exclusively detectable on brain microvascular endothelium of homozygous B-hLAT1 mice, but not in WT mice.
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Summary
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- mRNA expression analysis:
Mouse Slc7a5 mRNA was detectable only in brain of WT mice (+/+). Human SLC7A5 mRNA was detectable only in homozygous B-hLAT1 mice (H/H) but not in WT mice (+/+).
- Protein expression analysis:
LAT1 was detectable in WT mice (+/+) and homozygous B-hLAT1 mice (H/H) due to the cross-reactivity of antibodies. Human LAT1 was exclusively detectable in homozygous B-hLAT1 mice (H/H).
Human LAT1 was exclusively detectable on brain microvascular endothelium of homozygous B-hLAT1 mice, but not in WT mice.
