Analysis of HIV-1 viruses with a wild type PBS complementary to tRNALys,3 with mutated A-loop regions. Panel A. HIV-1 proviral genomes with the wild type PBS and A-loop (NL4-WT), wild type PBS and A-loop regions mutated to be complementary to tRNAHis (NL4-WT-HisAC) or tRNASer (NL4-WT-SerAC). Additional proviral constructs included HIV genomes with a PBS and A-loop region complementary to tRNAHis (NL4-His-HisAC). Panel B. Production of virus following transfection of wild type and mutant proviral genomes. 293T cells were transfected with equal amounts of proviral genomes and the amount of virus released into the supernatant was determined by p24 antigen capture ELISA. All viruses had similar amounts of p24 antigen into the culture supernatant (representative of three transfections). Panel C. Infectious virus was produced following transfection. Viruses were transfected with equal amounts of proviral genomes and the amount of infectious virus determined by the JC53-BL assay. Results are the average of the three determinations of infectivity in JC53-BL cells. Panel D. Replication of HIV-1 proviral genomes in SupT1 cells. Infections were established with equal amounts of infectious virus as determined by the JC53-BL assay. The amount of infectious virus was determined by the p24 antigen capture assay. The identity of the viruses are as marked.