Figure 5
Misincorporation assay. (A) Graphic representation of the template and primer system used to monitor the misincorporation efficiency of recombinant subtype B and C RT enzymes. The 32P-labeled 17-mer primer ppt17D annealed to 57-mer DNA template ppt57D was extended by HIV-1 subtype B and C recombinant RTs at 37°C for 5 min. The extension reactions were performed in the presence of all four complementary dNTPs, or, alternatively, in the absence of one of the dNTPs. The lanes marked with -A, -G, -C and -T indicate the missing nucleotide. Lanes marked with C indicate that all four dNTPs were included in the dNTP mix. Both RTs displayed similar levels of primer extension in the presence of all four dNTPs. P and FL indicate the positions of unextended primer and full-length extended products, respectively.