Development of soil amoeba Dictyostelium discoideum as an expression system for recombinant human erythropoietin

Abstract

Erythropoietin (EPO) is the most important biopharmaceutical product at present being produced by recombinant DNA technology from mammalian cell lines. Other available expression systems have not been useful in producing this protein due to the requirement of N-glycosylation for in vivo activity. In order to develop an alternative expression system, the human epo gene was expressed in the cellular slime mold Dictyostelium discoideum. The 2.43 kbp epo gene from the mammalian expression vector was cloned in the Dictyostelium expression cassette and used to transform cells. Positive clones were selected on the basis of antibiotic resistance. The clones were screened for the presence of the transgene. The copies of the gene inserted in the genome were identified and the transcript too was ascertained. The protein was identified by immuno-blotting and appears to be glycosylated though differently from that of humans or CHO cell lines. It has not yet been tested in bioassay.