Construction and in vitro characterization of a molecularly cloned human immunodeficiency virus type 1 library

Development of a safe and effective vaccine against human immunodeficiency virus (HIV) is urgent, but many concerns regarding the safety and efficacy of the currently developing vaccines remain. A major hindrance in HIV vaccine development is the genetic diversity, a hallmark of HIV biology, and a poor understanding of how HIV vaccine prevents the emergence of escape variants during infection and progression of AIDS. Here, we developed a method to construct a molecularly cloned viral library. This technique employs a long-range polymerase chain reaction (PCR) to amplify a virtually full-length HIV type 1 (HIV-1) provirus genome from peripheral blood mononuclear cells (PBMCs) infected with CRF01_AE (subtype E) Thai primary isolate. Among randomly selected 93 clones, 41 with a full-length sequence were able to replicate in PBMCs, 5 of which induced strong cytopathic effects. Replication kinetics also showed that the parental virus was intermediate among the clones. Thus, the molecular library prepared by this method showed the quasi-species in infected cells and this method could provide a new possibility for the development of an order-made therapeutic vaccine against HIV-1.