Sequence determination of the 3? end of mouse mammary tumor virus RNA

Abstract

70S RNA has been prepared from mouse mammary tumor virus (MMTV) produced by the GR tumor cell line. After denaturation for 3 min at 60 °C the RNA was applied to a sucrose gradient and molecules sedimenting between 10–16S were selected and passed over an oligo-dT cellulose column. The poly A containing RNA fraction was used as a template for the synthesis of complementary DNA with reverse transcriptase in the presence of dideoxynucleoside triphosphates. Oligo-nucleotides p(dT)7rG, p(dT)7rA and (p(dT)7rC were tested for their primer activity. Two of the three primers gave readable sequences. This suggests a heterogeneity in the 3′ end of the viral RNA, a phenomenon also observed with avian retroviruses. Nucleotides 37–45 from the 3′ end are made up of only A's and T's and resemble a Hogness box. This finding could have biological consequences for the transcription of proviral DNA in the unintegrated and integrated state. Instead of the usual AAUAAA sequence present in mRNA's close and in front of the site of polyadenylation we find a sequence AGUAAA.