Sequence characterization and expression analysis of mammary gland derived osteopontin gene of river buffalo

Abstract

Sequencing of RT-PCR amplified osteopontin (OPN) gene of buffalo mammary gland revealed its ORF to be of 843 nucleotides, coding for 280 amino acids long protein. Sequence comparison with cattle, sheep and pig showed 4 buffalo specific nucleotide changes. Two additional triplet nucleotides 273ACG275 and 681CAA683 in buffalo OPN as compared to cattle resulted in insertion of 2 amino acids N95 and N227. Phylogenetic analysis based on both nucleotides and amino acids sequence showed buffalo grouping more close to cattle and pig being placed most distantly from other 3 livestock species. Most of functional domains were conserved in buffalo OPN except that one of the 4 chymosin cleavage sites reported in cattle was missing in buffalo. Expression analysis of osteopontin gene by real-time PCR revealed approximately 5 fold increase in lactating buffalo mammary gland as compared to non lactating, indicating its importance in milk production.