Recombinant human granulocyte-macrophage-colony stimulating factor (GM-CSF)
US5891429A · kind A · utility
Assignee
Inventors
Key dates
| Filing date | Jun 6, 1995 |
| Grant date | Apr 6, 1999 |
| Priority date | — |
| Expiry date | Jun 6, 2015 |
Classification
- Technology area (CPC C)Chemistry; Metallurgy
- CPC primaryC12N2840/44
- WIPO fieldPharmaceuticals
- WIPO sectorChemistry
Abstract
A method for preparing and isolating a transformation vector containing CSF/cDNA is described. The method comprises: PA1 preparing RNA from a cell that produces CSF; PA1 preparing polyadenylated messenger RNA from said RNA; PA1 preparing single stranded cDNA from said messenger RNA; PA1 converting the single stranded cDNA to double stranded cDNA; PA1 inserting the double stranded cDNA into transformation vectors and transforming bacteria with said vector to form colonies; PA1 picking pools of 200 to 500 colonies each and isolating plasmid DNA from each pool; PA1 transfecting the plasmid DNA into suitable host cells for expressing CSF protein; PA1 culturing the transfected cells and assaying the supernatant for CSF activity; and PA1 selecting 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 t…
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