Reverse transcription of tRNA

Abstract

The 3′ terminus of tRNA was enzymatically elongated by an oligo(A) tail. A fragment of DNA polymerase I (E.coli) was used in the presence of manganese to phase and synthesize a cleavable primer at the oligo(A)-tRNA template. When the threedimensional structure of oligo(A)-tRNA is being unfolded under conditions where the primer is still hybridized at the oligo(A) tail, the DNA polymerase I fragment transcribes oligo(A)-tRNA into DNA. Reverse transcription is slowed down and its fidelity suspended by the 1-methyladenine in oligo(A)-tRNA^Phe(yeast). The reaction is stopped by the highly modified Y-base present in this template. Approximately full length transcripts can be obtained from oligo(A)-$tRNA3Gly$(E.coli). The transcription products were characterized by sequence analysis.