As a further step in the continuing process of defining the extent and nature of variability of the envelope (env) gene of HIV-1, we have cloned a new Zairian isolate, JY1, and sequenced the env gene of this isolate. Although the restriction map of the env region of JY1 was found to be more similar to that of the American prototype, BH10, than maps of all previously reported Zairian isolates and some American isolates, nucleotide sequencing of the JY1 env gene showed that it is among the most divergent from BH10 yet reported and that it differs from previously reported Zairian isolates almost to the same extent that it differs from BH10. A typical pattern of variable and constant regions was seen. A number of complex duplications were found in the hypervariable regions of JY1. The unique and highly divergent nature of the env gene of JY1 enhances its usefulness as part of a panel of HIV-1 isolates being evaluated in biologic and immunologic studies toward vaccine development.