Expression and characterization of the Mx2 protein in a heterologous expression system as a biomarker for early pregnancy detection in buffalo

The world’s largest producer of milk is India. This significant accomplishment is solely made possible by the existence of the Bubalus bubalis (Buffalo) species. The primary concern with buffaloes is the lack of early pregnancy detection, which is directly related to their high milk production. Thus, in the current investigation one of the con- ceptus-induced proteins, myxovirus resistance 2 (Mx2), which is produced between days 18 and 21 of pregnancy, was expressed and characterized in a heterologous protein production system. Partial Mx2 was markedly amplified in pregnant animals by conventional PCR. The suitability of the protein as a possible molecule for early pregnancy detection kits was determined using in silico analysis. The nature of the Mx2 protein under in vitro conditions was anticipated on the basis of its physiochemical properties, which are necessary for the formulation of any assay. Mx2 is highly antigenic in nature and can be exploited to produce antibodies, according to immunogenic characteristics. Using coding area primers, the partial Mx2 protein was extracted from a pregnant buffalo (n=30), amplified, and subsequently cloned and inserted into Escherichia coli BL21 (DE3). The 21 kDa band observed by western blotting showed that the protein is antigenic in nature. In conclusion, the expression and molecular characterization of partial Mx2 protein can be done in heterologous expression system. This Mx2 protein in buffalo may serve as a reliable marker of conceptus implantation, and can be incorporated into diagnostic tests for early pregnancy.