Simultaneous Expression of Multi-Subunit Proteins in Mammalian Cells Using a Convenient Set of Mammalian Cell Expression Vectors

摘要

To achieve fully functional heterolo-gous expression of multi-subunit re-combinant proteins, it is necessary to express several polypeptides within the cell at once. To address this requirement, we have developed a panel of four mammalian expression vectors in which all the components are identical except for the selectable markers. We present use of three of these vectors to express the alphal, alpha2 and (33 subunits of the human N-type calcium channel simultaneously in HEK293 cells (Catalog No. CRL 1.573; ATCC, Rockville, MD, USA). Optimum concentrations of each selection were determined, and cells were transfected by either calcium phosphate with supercoiled or linear plasmid DNA or by electroporation with linear plasmid DNA. After triple selection, transfectant colonies were screened for expression of each cDNA by reverse transcription polymerase chain reaction (RT-PCR). Transfection of linear plasmid DNA by electroporation yielded the highest percentage of transfectant cells expressing all three subunits.