Generation of VEGF knock-in Cashmere goat via the CRISPR/Cas9 system

Abstract

Cashmere is a rare and specialised animal fibre, which grows on the outer skin of goats. Owing its low yield and soft, light, and warm properties, it has a high economic value. Here, we attempted to improve existing cashmere goat breeds by simultaneously increasing their fibre length and cashmere yield. We attempted this by knocking in the vascular endothelial growth factor (<i>VEGF</i>) at the fibroblast growth factor 5(<i>FGF5</i>) site using a gene editing technology and then studying its hair growth-promoting mechanisms. We show that a combination of RS-1 and NU7441 significantly improve the efficiency of CRISPR/Cas9-mediated, homologous-directed repair without affecting the embryo cleavage rate or the percentages of embryos at different stages. In addition, we obtained a cashmere goat, which integrated the <i>VEGF</i> gene at the <i>FGF5</i> site, and the cashmere yield and fibre length of this gene-edited goat were improved. Through next-generation sequencing, we found that the up-regulation of <i>VEGF</i> and the down-regulation of <i>FGF5</i> affected the cell cycle, proliferation, and vascular tone through the PI3K-AKT signalling pathway and at extracellular matrix-receptor interactions. Owing to this, the gene-edited cashmere goat showed impressive cashmere performance. Overall, in this study, we generated a gene-edited cashmere goat by integrating <i>VEGF</i> at the <i>FGF5</i> site and provided an animal model for follow-up research on hair growth mechanisms.