Efficiency of tRNA
Lys-primed (-) ssDNA synthesis in cell-free assays. The efficiencies of synthesis of (-) ssDNA with HIV-1 subtype B and subtype C wild-type RTs were compared in time course experiments using 5'-end 32P-labeled human natural tRNA3Lys as primer and HIV-1 PBS RNA as template. The HIV-1 PBS RNA template used in this system consists of 258 nucleotides (nt) at the 5' end of the HIV-1 genome, which contains the R, U5, and PBS regions. Synthesis of full-length DNA (FL DNA) by recombinant RT enzymes was monitored in time-course experiments. Reactions were initiated by the addition of MgCl2 and dNTPs and stopped at different time points during a period of 45 min. The position of the full-length DNA product (FL DNA) is shown on the right.