The process of making a recombinant protein is simple. First, a gene encoding the desired protein is isolated and sequenced. This gene is then inserted into an appropriate expression vector, which directs the gene to be expressed in a host organism. The recombinant protein can then be produced by culturing the host organism with the expression vector.
The culturing step can be performed in large culture vessels containing whole bacteria or cells, or in a fermenter or bioreactor vessel. The recombinant protein is harvested from the culture medium and purified by conventional techniques such as preparative high performance liquid chromatography (HPLC) and electrophoresis.
A final purification step may be necessary to further purify the protein if it contains contaminants such as nucleic acid sequences. Methods for the production of Recombinant Proteins are well known in the art, see e.g., Sambrook et al.,
Molecular Cloning: A Laboratory Manual 2nd Edition, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y., 1989; Kriegler et al.,
Gene Transfer and Expression: A Laboratory Manual, Stockton Press, New York, 1989; and the recombinant DNA manuals of Ausubel et al., Current Protocols in Molecular Biology (1991 Supplement) John Wiley & Sons, Inc. N.Y.; and Beigelman et al., Current Protocol in Immunology (1994 Supplement) J. E. Coligan et al., Eds Greene Publishing and Wiley-Interscience Publishers, 1994; all three of which are incorporated herein by reference for any purpose.
The Recombinant Proteins can be produced in vitro by a variety of methods that are well known in the art including chemical synthesis, fermentation methods and recombinant DNA technology methods as described above or any combination thereof. The recombinant proteins disclosed herein can also be produced in recombinant cells such as bacteria and yeast.