Chemical synthesis and improved expression of recombinant human granulocyte colony-stimulating factor cDNA

Abstract

Recently, granulocyte colony-stimulating factor (G-CSF) has been recognized as a useful molecule for the treatment of a wide range of complex ailments, such as cancer, AIDS, H1N1 influenza, cardiac and neurological diseases. The vast therapeutic potential of G-CSF has induced scientists to develop biotechnological approaches for the synthesis of this pharmacologically active agent. We used a synthetic G-CSF cDNA molecule to produce the target protein by a simple cloning protocol. We constructed the synthetic cDNA using a DNA synthesizer with the aim to increase its expression level by specific sequence modifications at the 5' end of the G-CSF-coding region, decreasing the GC content without altering the predicted amino acid sequences. The identity of the resulting protein was confirmed by a highly specific enzyme-linked immunosorbent assay. In conclusion, a synthetic G-CSF cDNA in combina tion with the recombinant DNA protocol offers a rapid and reliable strategy for synthesizing the target protein. However, commercial utilization of this methodology will require rigorous validation and quality control.