U5-PBS sequence and Infectivity levels of HIV-1 NL4-3 viral mutants with altered U5 and PBS sequences at start of PBMC infection. Panel A. HIV-1 U5 and PBS sequence (shown 5' to 3'). Viral primer binding site (PBS) and U5 A-loop sequences were altered to be complementary to the 3' terminal 18 nucleotides and anticodon loop of tRNAIle, tRNAMet, tRNAPro, and tRNATrp, respectively. U5 A-loop sequence and PBS are shadowed. Panel B. Comparison of the infectivity of U5-PBS mutant NL4-3 viruses. HIV-1 NL4-3 proviral clones were transfected into 293T cells, incubated for 48 hours, and supernatants were measured for infectious units. For a given sample, the number of infectious units per microliter is equal to the number of blue cells in a well divided by the dilution factor for that well and represents the average of at least two wells. Wild type infectivity levels were set at 100%, and mutant virus infectivity was reported as a percentage of wild type. All viruses with altered U5 and PBS sequences had reduced levels of infectivity as compared to wild type virus. The data presented are representative for three independent experiments.