Human immunodeficiency virus type 1 (HIV-1) and type 2 (HIV-2) differ in their pathogenic mechanisms as evidenced by lower rate of disease progression, lower transmission rates and lower viral load in peripheral blood for HIV-2. One of the many factors that are involved in these characteristics is the interaction between viral glycoproteins and cellular receptors. The study of these interactions in an HIV-2 model could lead to important conclusions regarding pathogenesis and transmission mechanisms of HIV-2 infection. Here we report the design of a method enabling the construction of recombinant proviral HIV-2 DNAs in a moderate copy number plasmid that allows the analysis of env gene structure and functionality. This method constitutes an important tool for the study of HIV-2 env glycoproteins and for the mappings of genetic determinants of HIV-2 coreceptor usage and CD4-independent interaction. Furthermore, this knowledge will help towards the understanding of the different pathogenic mechanisms of HIV-1 and HIV-2.