Animal Molecular Breeding 2024, Vol.14, No.1, 106-118 http://animalscipublisher.com/index.php/amb 111 Figure 3 Generation of GGTA1−/−β2M−/−CIITA−/−triple gene knockout (GBC-3KO) pigs (Adopted from Fu et al., 2020) Image caption: A, Schematic overview of the generation of GBC-3KO pigs. B, Illustration of the CRISPR/Cas9 targeting sites in GGTA1, β2M, and CIITA genes. The sgRNA targeting sequence is underlined in black, and the protospacer-adjacent motif (PAM) sequence is underlined and labeled in red. C, Photograph of GBC-3KO pigs from GBC-21 porcine embryonic fibroblast cell line. D, Genotyping of the GBC-3KO pigs by polymerase chain reaction (PCR). E, Genotyping of the GBC-3KO pigs from GBC-21 porcine embryonic fibroblast cell line by Sanger sequencing. The sizes of insertion (+) and deletion (Δ) are presented on the right side of each allele. β2M, β2-microglobulin; Cas9-eGFP, pUC19-pCAG-SpCas9-2A-GFP; CIITA, major histocompatibility complex class II transactivator; CRISPR/Cas9, a gene-editing technology; GGTA1, glycoprotein galactosyltransferase α 1, 3; sgRNA, single guide RNA sequence 7; PEF, pig embryonic fibroblast; U6-gRNA, pUC19-U6-sgRNA; WT, wild type (Adopted from Fu et al., 2020)
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