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Fig. 1 | Acta Neuropathologica Communications

Fig. 1

From: CMT2Q-causing mutation in the Dhtkd1 gene lead to sensory defects, mitochondrial accumulation and altered metabolism in a knock-in mouse model

Fig. 1

Strategy used for establishing the Dhtkd1Y486* knock-in mouse model and the expression profile of Dhtkd1. a Schematic illustration of the strategy used to generate the Dhtkd1Y486* knock-in mice. The first schematic line indicates the Dhtkd1 allele in the wild-type (wt) mouse. Exons and introns are represented by boxes and horizontal lines, respectively. The coding regions are shown in black. Positions of the start codon and the stop codon are indicated by star and delta sign, respectively. Homologous recombination results in the point mutation and the replacement of a small part of the intron between exon 8 and exon 9. The second line shows the targeting construct. PGK-Neo cassettes are used for positive selections. The third line shows the targeting alleles. P1-P8 indicates the primers used for genotyping. b Identification of positive ES cell clones by PCR amplification with primers indicated in the Fig. M = DNA molecular ladder. c Sequencing of positive ES cell clones confirms the mutation, with the DNA base variations indicated in the figure by arrows. d Relative Dhtkd1 mRNA level, normalized to Gapdh, from ES cells showing a significant decrease in Dhtkd1 mRNA level in mutant lines. e Genotyping showed the Dhtkd1 in wt, heterozygous (wt/mt), and homozygous (mt/mt) mice with respect to primers P7-P2 and P7-P8. f Sequencing of genomic DNA from mouse tails confirms the mutation. Sequence DNA base variations are indicated by arrows. g RT-PCR of Dhtkd1 starting from total RNA from liver and sciatic nerve in wt, wt/mt, and mt/mt mice. Gapdh was used as an internal control. h Western blot with kidney tissue of wt, wt/mt, and mt/mt mice. One of three independent experiments in triplicate is shown. ***p < 0.001

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