A bicistronic expression vector, p3.1-IRES-CD34, has been constructed to facilitate the selection and screening of transfected cells by magnetic cell sorting (MACS). In this vector, an engineered variant of CD34, deleted CD34 (dCD34), was chosen as the marker gene and the internal ribosome entry site (IRES) from encephalomyocarditis virus (EMCV) was employed to allow simultaneous expression of the inserted 5′-end heterologous gene and the CD34 marker gene. To test the utility of the vector, the enhanced green fluorescent protein (EGFP) gene was inserted into the multiple cloning site (MCS) of the vector. Transfected cells were selected by magnetic cell sorting (MACS). The results demonstrated that the transfected cells can be enriched to high purity (>95%). The vector p3.1-IRES-CD34 would provide a rapid (2~3 days for transient transfected cells and 10 ~ 15 days for stable transfected cells) and efficient method for the selection of transfected cells.