The project outlined in this proposal is designed to complement earlier research I have performed on the biology of the replication of the human immunodeficiency virus, type 1 (HIV-1) with training in the techniques of molecular biology. My goal is to learn to apply these techniques to answer questions regarding the encapsidation of the viral genomic RNA. Specifically, I plan to subject both the putative RNA packaging signal as well as the nucleocapsid protein to random mutagenesis. Mutants will be pooled and selected for viability simply by their ability to grow in culture. Their genomes will be recovered by PCR amplification of infected cells. Sequence information will point to either nucleotides, in the case of the RNA packaging signal, or amino acids that are intolerant of substitution. In this way I hope to identify those residues which play important structural roles in RNA packaging. Given the impact of HIV infection and the crucial role RNA packaging plays in the formation of infectious particles, an understanding of the mechanisms by which HIV recognizes its genomic RNA is an important health-related aspect of this work.
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