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jbc

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jbc

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  1. Supplemental Figure. Extension by HIV-RT with different divalent cations on an unstructured template. 5‘-32P-radiolabeled oligo-dT20 primer was hybridized 1:10 (w:w) to poly-rA template and added at a final concentration of 0.002 μg/μl in rA to a reaction mixture containing 100 μM dTTP, 50 mM Tris-HCl (pH=8), 80 mM KCl, 1 mM DTT, and either 2 mM MgCl2, 400 μM ZnCl2, or both. To initiate reactions, RT was added at a final concentration of 25 nM. Aliquots were removed and extension stopped with 2X formamide-dye at 5,15, 30 seconds, 1, and 2 minutes for MgCl2-only reactions or 5, 15, 30 seconds, 1, 2, 4, 8, and 16 minutes for reactions containing ZnCl2. The maximum extension rate was ~25 times faster in the presence of MgCl2 than ZnCl2 (~25 nt/sec and ~1 nt/sec, respectively). Extension in the presence of both cations is indistinguishable from ZnCl2-only reactions. Lane M, size marker in nucleotides; land –E, no enzyme added.

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