Synthesis and characterization of DNA complementary to carnation mottle virus RNA.
DNA complementary to carnation mottle virus-RNA (CarMV RNA) was synthesized using the reverse transcriptase of avian myeloblastosis virus (AMV) as enzyme, and an hydrolysate of calf thymus DNA as primer. After purification steps that included self-annealing and hydroxyapatite chromatography, the DNA transcript was more than 99% sensitive to S1 nuclease and hybridized to at least 92% of the CarMV-RNA sequences, but not to heterologous RNA. Therefore, the purified transcript was single stranded and represented almost all CarMV-RNA sequences. The cDNA consisted of small pieces that hybridized to CarMV-RNA with a sharp transition corresponding to Crt(1/2) of 2.3 x 10(2) M sec liter(-1). This value is consistent with the complexity expected for CarMV-RNA if there is only one kind of virion RNA. CarMV-cDNA hybridized completely to a crude RNA preparation from CarMV-infected carnation leaves, but not to a comparable RNA preparation from healthy leaves, thus permitting the use of CarMV-cDNA to detect and to quantify the viral nucleotide sequences in total nucleic acid preparations from fragments of carnation cuttings.
Kummert
J
J
Laboratoire de Pathologie Végétale, Faculté des Sciences Agronomiques, 5800 Gembloux, Belgium.