To show the flexible adapter design of restriction display (RD) technique in constructing peptide library, two series of adapters were designed according to vector pET22b in E.coli and vector pNMT-TOPO in S.pombe. Each series of adapter has three in turn one-base-increment adapters, which allowed the inserted DNA fragment probably expressed in the correct reading frame. HIV-1 subtype B whole gene was used as an example, and fragments expression libraries with two different adapters were constructed by RD technique, randomly 12 clones from each library were sequenced for translation analysis. As a result, a clone from the prokaryotes library was obtained, which could encode HIV Pol peptide. Then the positive plasmid was induced to express protein in E.coli BL21(DE3). SDS-PAGE and Western blot showed positive result, which is consistent with expected. It can be concluded that RD technique is a new approach for constructing genome random peptide library, and its flexible adapters design can meet with kinds of expression vectors.