Cloning, characterization and expression of bovine (<i style="">Bos indicus</i>) tumour necrosis factor-<img src='/image/spc_char/alpha.gif' border=0>

Abstract

The gene for tumour necrosis factor-alpha (TNF-<img src='/image/spc_char/alpha.gif' border=0> ) was amplified from cDNA pool prepared from LPS-stimulated bovine peripheral blood mononuclear cells isolated from Indian cattle. The amplified TNF-<img src='/image/spc_char/alpha.gif' border=0> gene was cloned and nucleotide sequences were determined. Homology comparison of nucleotide and predicted amino acid sequences revealed similarity at nucleotide and amino acid level in both exotic cattle and goat. The coding sequence of mature TNF-<img src='/image/spc_char/alpha.gif' border=0> (without its signal sequence and transmembrane anchor) was expressed as fusion protein with N-terminal polyhistidine using prokaryotic expression vector. The expressed protein was present as insoluble inclusion bodies. The recombinant protein was solubilized with urea and purified using Ni-agarose affinity chromatography. The purified recombinant TNF-<img src='/image/spc_char/alpha.gif' border=0> was characterized in SDS-PAGE and in western blotting.