Antibody production in Chinese hamster ovary cells using an impaired selectable marker

The expression levels of transfected genes in mammalian cells are primarily determined by the cellular DNA at the site of integration. In this report we describe immunoglobulin expression vectors designed to target mammalian loci that support high levels of expression. These vectors encode immunoglobulin light and heavy chain genes, the dihydrofolate reductase (DHFR) gene, and the dominant selectable marker neomycin phosphotransferase (NEO) gene. Selectivity of the vectors has been achieved by intentional impairment of the NEO gene by two different mechanisms. The NEO translation initiation site has been impaired and an intron has been introduced into the gene. Selection in G418 for clones containing the NEO gene yields an increased percentage of high expression clones from a decreased number of total clones. The majority of high expressing clones contain a single gene copy. Subsequent gene amplification results in clones producing very high levels of immunoglobulin with only a few gene copies.