FISSION YEAST EXPRESSION VECTORS ADAPTED FOR POSITIVE IDENTIFICATION OF GENE INSERTION AND GREEN FLUORESCENT PROTEIN FUSION

A pm series of fission yeast expression vectors, derivatives of the pR EP series, was designed to allow positive identification of cloned gen e insertion and fusion to the green fluorescent protein (GFP) gene for in vivo analysis of gene expression. To validate this new vector syst em, the human immunodeficiency virus type 1 (HIV-1) vpr gene of viral isolate pNL4-3 was expressed in the pYZ1N vector: Vpr-induced phenotyp ic changes were the same as those observed with vpr expressed from pRE P1N. Consistent with observations in mammalian cells, a Vpr-GFP fusion protein localizes on the nuclear membrane of fission yeast cells. Add itionally, we were able to detect a naturally occurring mixture of vpr genes from a plasma sample of an HIV-infected pediatric long-term sur viving patient. These pYZ vectors expedite gene cloning for general pu rposes and are particularly suited for large-scale random gene screeni ng.