Synthesis in High Yield of Complementary DNA of Retroviral RNA

Abstract

Mouse mammary tumor virus RNA was transcribed in vitro with avian myeloblastosis virus reverse transcriptase in the presence of calf thymus DNA oligomers. The yield of cDNA was markedly enhanced by increasing the concentration of the enzyme as well as primers in the reaction mixture. The average size of cDNA was approximately 200 residues, and it was not affected by the concentration of deoxynucleoside triphosphates when saturating concentration of enzyme was used. Nearly all of the viral sequences were represented in cDNA and it formed hybrids of high fidelity with viral RNA. Similar results were obtained when murine leukemia virus (AKR) RNA was transcribed. These observations will be useful for synthesizing cDNA of RNAs that are not easily available in sufficient quantities.