Generation of conditional Acvrl1 knockout mice by CRISPR/Cas9-mediated gene targeting

Abstract

Objectives: This study aimed to generate mutant mice containing the Acvrl1 gene flanked with LoxP sequences to allow conditional deletion of Acvrl1 by the LoxP/Cre system. Such mice may facilitate the development of brain arteriovenous malformation (BAVM) models.

Methods: The CRISPR/Cas9 technique was used to edit Acvrl1. Two single guide RNAs (sgRNAs) with recognition sites on intron 3 and 8 and a donor vector that was homologous with the targeted gene and contained two LoxP sequences were designed and constructed. The in vitro-synthesized sgRNA, Cas9 mRNA and donor vectors were injected into mouse zygotes, which were then transferred into pseudopregnant mice. Neonatal mutant mice were identified by genotyping and sequencing.

Results: Two mice with a floxed Acvrl1 allele were generated at a success rate of 8.7%. The target mice, which were healthy and fertile, were obtained through interbreeding.

Conclusion: CRISPR/Cas9 is a reliable gene-editing tool, and is able to efficiently modify Acvrl1 and create the target mice.

Keywords: Acvrl1; Arteriovenous malformation; CRISPR/Cas9; Mouse.