We have demonstrated that the synthetic oligonucleotides, either unmodified or phosphorothioates, prevent cDNA synthesis by the AMV or HIV reverse transcriptases. The RNA was truncated at the antisense oligonucleotide-RNA duplex during the reverse transcription. Th blockage involves the degradation of the RNA fragment bound to the antisense oligonucleotide by the reverse transcriptase associated RNase H activity. However, in the case of phosphorothioate oligomer, the production of cDNA would be inhibited by a hybrid formed between the AMV RT and phosphorothioate oligomer than arrested elongation of the cDNA strands, whereas arrest of a growing cDNA strand by HIV RT can be blocked by an oligonucleotide complementary to a region downstream from the primer.