Recombinant glutathione S-transferase (GST) Vpr fusion protein was used as target to select Vpr binding peptide from phage peptide library. By immobilized the GST or GST fusion protein to glutathione agarose gel, binding phage could be quickly panned and eluted by reduced glutathione. The sequencing result show that a WWXF motif exist among the Vpr binding peptide. Comparing with the classic biopanning method by coating target protein on plate this provide more quick and convenient way to screen binding protein from phage peptide library.