Sequencing minus-strand strong-stop DNA ([−] ssDNA) to identify sequence of HTLV-1 and HIV-1 RT primers. (A) Flowchart of method used to amplify sequence containing the HIV-1 and HTLV-1 RT primer with a short RT extension. Endogenous
RT was performed to synthesize the (−) ssDNA (step 1). Following demethylation, a second round of RT was performed to synthesize
the antisense strand of the tRNA RT primer (step 2). The product was PCR-amplified (steps 3 and 4). (B,C) The PCR-amplified products were run on 2% agarose gels. PCR products with the expected size (boxed with red lines, 135 bp
for HTLV-1, B; 133 bp for HIV-1, C) were excised and eluted from the gel and submitted for Sanger sequencing. (D,E) Chromatograph of Sanger sequencing results for the (−) ssDNA containing RT primer and sequence alignments to 
(for HTLV-1, D) or 
(for HIV-1, E).
