Recombinant human granulocyte-macrophage-colony stimulating factor (GM- CSF)

摘要

A method for preparing and isolating a transformation vector containing CSF/cDNA is described. The method comprises:PP preparing RNA from a cell that produces CSF;PPpreparing polyadenylated messenger RNA from said RNA;PPpreparing single stranded cDNA from said messenger RNA;PPconverting the single stranded cDNA to double stranded cDNA;PP inserting the double stranded cDNA into transformation vectors and transforming bacteria with said vector to form colonies;PP picking pools of 200 to 500 colonies each and isolating plasmid DNA from each pool;PPtransfecting the plasmid DNA into suitable host cells for expressing CSF protein;PPculturing the transfected cells and assaying the supernatant for CSF activity; and P Pselecting CSF positive pools and screening the colonies used to make the pool to identify a colony having CSF activity. Also described are a cDNA coding for a protein having CSF activity (i.e. CSF/cDNA), a microorganism or cell line transformed with a recombinant vector containing such CSF/cDNA, and a method for producing CSF protein by expressing said CSF/cDNA by culturing a microorganism or cell line. The invention also provides a method of purifying the CSF proteins and the purified proteins so produced.