Expression Of Human Recombinant Tumor Necrosis Factor(TNF) And The Fusion Of IFN-γ And TNF Proteins Using Oligonucleotde-Directed Deletion-Insertion Mutagenesis

Tumor necrosis factor(TNF), a factor mainly secreted by macrophages ,was first reported in 1975 by Carswell et al that the serum of endotoxin treated mice, rats, and rabbits, previously infected with Mycobacterium bovis strain Bacillus Calmette-Gunerin, caused hemorrhagic necrosis of various tumor in mice with no apparent effect on the host. TNF also displays cytostatic and cytotoxic activities against several transformed cell lines in culture, but normal cell cultures derived from human, rabbit, chicken, and mouse souces are unaffected. No normal cells have yet been described as seneitive to TNF so far.Besides tumoricidal activity, TNF is also found to be an antiviral reagent, which can inhibit the replication of various viruses, and pretect the cultured cells from viral infertion. Recent studies showed that TNF is potential as a immunomodutator,activating several immunoeffective cells as macrophages, neutrophils , and T and B lymphocytes, inducing the expression of major histocompatible complex antigen, and enhancing the function of immune system against different pathogenic microbes invaded.Nevertheless, the studies on the functions, structure and clinical applications of TNF is not extensively carried out inland becaues of the limitation of TNF resources, which is not abundant in nature, and the production of tumor necrosis factor by the genetic engineering method seems to be the only way to solve this problem.The present paper described the research work about the expression of the tumor necrosis factor,in which we deleted the noncoding sequence and the leading peptide - coding sequence upstream the tumor necrosis factor gene, and inserted a translational initiation codon ATG in front of the first amino acid codon of TNF, by utilising the oligonucleotide "directed mutagenesis. The high level of expression of tumor necrosis...