Generation of PECAM‐1 (CD31) conditional knockout mice

Abstract

Summary Platelet endothelial cell adhesion molecule 1 (PECAM‐1) is an adhesion and signaling receptor that is expressed on endothelial and hematopoietic cells and plays important roles in angiogenesis, vascular permeability, and regulation of cellular responsiveness. To better understanding the tissue specificity of PECAM‐1 functions, we generated mice in which PECAM1, the gene encoding PECAM‐1, could be conditionally knocked out. A targeting construct was created that contains loxP sites flanking PECAM1 exons 1 and 2 and a neomycin resistance gene flanked by flippase recognition target (FRT) sites that was positioned upstream of the 3′ loxP site. The targeting construct was electroporated into C57BL/6 embryonic stem (ES) cells, and correctly targeted ES cells were injected into C57BL/6 blastocysts, which were implanted into pseudo‐pregnant females. Resulting chimeric animals were bred with transgenic mice expressing Flippase 1 (FLP1) to remove the FRT‐flanked neomycin resistance gene and mice heterozygous for the floxed PECAM1 allele were bred with each other to obtain homozygous PECAM1 flox/flox offspring, which expressed PECAM‐1 at normal levels and had no overt phenotype. PECAM1 flox/flox mice were bred with mice expressing Cre recombinase under the control of the SRY‐box containing gene 2 (Sox2Cre) promoter to delete the floxed PECAM1 allele in offspring (Sox2Cre;PECAM1 del/WT), which were crossbred to generate Sox2Cre; PECAM1 del/del offspring. Sox2Cre; PECAM1 del/del mice recapitulated the phenotype of conventional global PECAM‐1 knockout mice. PECAM1 flox/flox mice will be useful for studying distinct roles of PECAM‐1 in tissue specific contexts and to gain insights into the roles that PECAM‐1 plays in blood and vascular cell function.

Development and characterization of an Otp conditional loss of function allele

“…To generate a conditional Otp knockout allele, we inserted LoxP sites to flank exon 2 of the mouse Otp gene using the CRISPR‐Cas9 system (Hai, Szwarc, Lanza, Heaney, & Lydon, 2019; Zhi, Kanaji, Fu, Newman, & Newman, 2020). To this end, we first designed and validated sgRNAs targeting the intronic sequences flanking exon 2 of the mouse Otp gene.…”

Conditional knockout mouse model reveals a critical role of peroxiredoxin 1 in oral leukoplakia carcinogenesis

“…To achieve this, we employed CRISPR/Cas9 technology and utilized K14-cre mice to selectively knock out the Prx1 gene within the tongue epithelium of mice. The utilization of different Cre mouse models allows for conditional gene knockout in specific tissues, facilitating the investigation of gene function in a tissue-specific context [ 30 , 31 ]. In K14-Cre mice, the Cre recombinase is produced by the regulation of the human K14 promoter [ 32 ].…”

Transcriptomic analysis reveals that bromodomain containing 9 controls signaling pathways in gastric cancer

“…It is expressed on endothelial cells and hematopoietic cells and regulates angiogenesis, vascular permeability, and cell reactivity [18]. PECAM-1 plays an important role in interference with tumor clearance [19][20][21][22], and Terashima et al found that it is substantially related to peritoneal recurrence in patients with stage II and III gastric cancer. This molecule is involved in several processes related to the growth and spread of primary cancer by encoding proteins, including angiogenesis, vascular permeability, and extracellular leukocyte transport.…”