Isolation of genomic DNA controlling mouse melanoma antigen defined by monoclonal antibody

We have isolated the genomic DNA controlling the expression of murine specific melanoma antigen by employing cosmid shuttle vector and monoclonal antibody. Transfection of the cosmid library derived from mouse melanoma cells into human melanomas and repeated cell sortings of the fluorescence-bright population enabled us to enrich the antigen-positive transfectants. We rescued a 34.8 kb DNA fragment from the transfectants by in vitro packaging and showed it to be responsible for the antigen expression. However, we noticed instability of the antigen expression when the selection pressure imposed by the cell sorting was removed. This seemed to be due to the fact that the insert DNA was preferentially deleted from this cosmid vector without loss of the vector sequence itself.