Method for identification and isolation of DNA encoding a desired protein

摘要

A method for identifying and isolating clones containing DNA coding for a desired protein is described. DNA prepared from a cell that expresses the desired protein is inserted into an isolation expression vector having means for replication (as a means of producing DNA) and a suitable promoter for expression of said DNA in a predetermined mammalian host cell as well as means for replication in a bacterial cell. The transient expression vector is then inserted into a bacterial cell for replication of the DNA. Pools of DNA, prepared from a predetermined number of bacterial clones so that the nucleic acids (DNA and RNA) is substantially free of other bacterial contaminants are transfected or microinjected into mammalian host cells and conditioned medium from growing such cells is tested for the presence of the desired protein. Positive pools are selected and the clones used to make the pool are screened to identify and isolate the clone containing the desired DNA.