Efficient N-Tailing of blunt DNA ends by Moloney murine leukemia virus reverse transcriptase

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    Abstract

    Moloney murine leukemia virus reverse transcriptase (MMLV-RT) is a widely used enzyme for cDNA synthesis. Here we show that MMLV-RT has a strong template-independent polymerase activity using blunt DNA ends as substrate that generates 3′ overhangs of A, C, G, or T. Nucleotides were appended efficiently in the order A > G > T > C, and tail lengths varied from 4 to 5, 2 to 7, 2 to 4, and 2 to 3 for A, C, G, and T, respectively. The activity was so strong that nearly all our test DNA ends were appended with at least one A, C, G, or T. The N-Tailing activity of MMLV-RT was enhanced in the presence of Mn 2+, and the G-, C-, and T-Tailing activities were further enhanced by dCMP, dGMP, and dAMP, respectively. This is the first report of an enzymatic activity that almost thoroughly appends two or more As, or one or more Cs, Gs, or Ts to the 3′ end of double-stranded DNA, which would enable exhaustive analysis of DNA samples. The N-Tailing activity of MMLV-RT is potentially useful in many biotechnological applications.

    Original languageEnglish
    Article number41769
    JournalScientific reports
    Volume7
    DOIs
    Publication statusPublished - 2017 Feb 2

    ASJC Scopus subject areas

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