Generation of prolactin-inducible protein (Pip) knockout mice by CRISPR/Cas9-mediated gene engineering

• Published in: Canadian journal of physiology and pharmacology Vol. 100; no. 1; pp. 86 - 91
• Main Authors: Terceiro, Lucas E.L, Blanchard, Anne A.A, Edechi, Chidalu A, Freznosa, Agnes, Triggs-Raine, Barbara, Leygue, Etienne, Myal, Yvonne
• Format: Journal Article
• Language: English
• Published: Canada Canadian Science Publishing 01.01.2022; NRC Research Press; Canadian Science Publishing NRC Research Press
• Subjects: Animals; Breast cancer; CRISPR; CRISPR-Associated Protein 9 - genetics; CRISPR-Cas Systems - genetics; CRISPR/Cas9; Embryos; Exons; Gene Knockout Techniques - methods; Genetic diversity; Genetic Engineering - methods; Genetically modified mice; Genotyping; Glycoproteins; Growth; Health aspects; Inbreeding; Keratoconus; Mammary gland; Mice; Mice, Inbred C57BL; Mice, Knockout; Models, Animal; modèle de souris transplantable; Physiological aspects; Prolactin; Prolactin-inducible protein; Protein engineering; Proteins; Proteins - genetics; protéine inductible par la prolactine; Rodents; Secretions; Sjogren's syndrome; Sweat gland; transplantable mouse model; Canada; transplantable mouse model; CRISPR/Cas9; protéine inductible par la prolactine; prolactin-inducible protein; modèle de souris transplantable
• ISSN: 0008-4212; 1205-7541
• DOI: 10.1139/cjpp-2021-0306

Prolactin-inducible protein (PIP) is a multifunctional glycoprotein that is highly expressed and found in the secretions of apocrine glands such as salivary, lacrimal, and sweat glands including the mammary glands. PIP has been implicated in various diseases, including breast cancer, gross cystic disease of the breast, keratoconus of the eye, and the autoimmune Sjögren’s syndrome. Here we have generated a Pip knockout (KO) mouse using the Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/CRSPR-associated (Cas)9 system. The Cas9 protein and two single guide RNAs targeting specific regions for both exons 1 and 2 of the Pip gene were microinjected into mouse embryos. The deletions and insertions promoted by CRISPR/Cas9 system on the Pip gene successfully disrupted Pip protein coding, as confirmed by PCR genotyping, sequencing, and ultimately Western blot analysis. This mouse model was generated in the inbred C57Bl/6J mouse, which exhibits lower genetic variation. This novel CRISPR Pip KO mouse model will not only be useful for future studies to interrogate the multifunctional role of PIP in physiological processes but will facilitate a broader understanding of the function of PIP in vivo while providing unprecedented insight into its role in a spectrum of diseases attributed to the deregulation of the PIP gene.