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. 2015 Oct 16;16(10):24732–24750. doi: 10.3390/ijms161024732

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© 2015 by the authors; licensee MDPI, Basel, Switzerland.

This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution license (http://creativecommons.org/licenses/by/4.0/).

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CRISPR/Cas9 strategy and results for Ccdc63^+1/+1 male mice. (A) Ccdc63 exon 4 was targeted; coding exons shown as solid black bars, non-coding exons shown as open bars. Forward (Fw) and reverse (Rv) primers for validation and genotyping shown; black arrows labelled Fw and Rv. sgRNA sequence underlined in black. PAM sequence is shown in red. Blue arrowhead indicates site of Cas9 digestion; (B) Sequencing of Ccdc63^+1/+1 mice; black arrow and bar indicates the nucleotide inserted; (C) Frameshift mutation of Ccdc63^+1/+1, mutation shown in red, * indicates a premature stop codon. Amino acid number shown above sequence; (D) Natural mating analysis of Ccdc63^+1/+1 male mice with WT females. Pregnancy rate (pregnancy/vaginal plug) presented as a percentage.