Extended Data Fig. 10: Generation of a PCDH1-KO knockout Syrian hamster by CRISPR–Cas9 genome engineering. | Nature

Extended Data Fig. 10: Generation of a PCDH1-KO knockout Syrian hamster by CRISPR–Cas9 genome engineering.

From: Protocadherin-1 is essential for cell entry by New World hantaviruses

Extended Data Fig. 10

a, Organization of the PCDH1 gene of the Syrian hamster (M. auratus). The sequence in exon 2 of PCDH1 that was targeted by an sgRNA is shown in magenta. Knockout animals bear two PCDH1 alleles that have been edited to lack a single nucleotide, highlighted in green. The sgRNA PAM is highlighted in blue. b, A PCR–RFLP strategy, based on loss of digestion by the restriction endonuclease BanI, was used to detect genome editing and genotype animals. c, PCR–RFLP results were confirmed by Sanger DNA sequencing of PCR amplicons from WT and genome-edited animals. Sequencing traces are shown. Sequence features are highlighted as in a. Red arrows denote the site of gene editing in the PCDH1-KO allele. Experiments were performed twice with similar results. d, Lung tissue isolated from WT and PCDH1-KO hamsters was solubilized, normalized by protein content, and subjected to SDS–PAGE. PCDH1 was detected by immunoblotting with EC1-specific mAb 3305. Control, nonspecific loading control. Experiments were performed three times with similar results. e, Viral loads in the sera of WT and PCDH1-KO hamsters at 14 dpi. The limit of detection is shown as a dotted line. Experiments in be were performed twice with similar results.