Isolation, bioinformatic analysis and tissue expression profile of a novel water buffalo gene-<i>MFG-E8</i>

Abstract

Abstract. Milk fat globule-epidermal growth factor 8 (MFG-E8) is a milk membrane-associated glycoprotein, which plays a critical role in phagocytic clearance of apoptotic cells and mammary gland development, involution and remodeling. In the present study, the complete CDS of water buffalo MFG-E8 was obtained and characterized. The genetic variations of nine water buffalo were investigated. Further, the tissue expression profile was carried out using Quantitative real-time PCR method. The full-length coding region of MFG-E8 from water buffalo tissues consists of 1296 nucleotides, which encodes 431 amino acids with a molecular weight of 47.85 kDa and a pI of 7.02. Bioinformatic prediction indicates that the MFG-E8 protein contains one signal peptide, two repeated EGF-like domains in its N-terminal side and two repeated discoidinlike Factor 5/Factor 8 domains in its amino acids. The sequence homology analysis in Bovidae family revealed that the coding region of water buffalo MFG-E8 had 98.5, 98.3, 93.9, and 88.0 % identity with that of cattle, yak, goat and sheep. There was no polymorphism found in water buffalo, but there existed five nucleotide differences between water buffalo and other bovine species. The phylogenetic tree based on the amino acid sequences of MFG-E8 from thirteen species revealed that buffalo had a closer genetic relationship with the species of Bovidae family. Real-time PCR analysis showed that water buffalo MFG-E8 gene was expressed in various tissues, but at different levels. The expression levels of this gene were higher in the mammary gland than in other tissues, suggesting that the MFG-E8 protein plays a role in mammary gland functions.