Reverse Transcription by Escherichia coli DNA Polymerase I

Abstract

E. coli DNA polymerase I (EC 2.7.7.7) can engage in either DNA- or RNA-directed DNA synthesis with hybrid templates. The choice of the strand to be transcribed depends primarily on the relative lengths of the two strands of the hybrid, the longer strand serving as the template and the shorter as the primer. If a polynucleotide is reduced in size by exposure to an endonuclease before being hybridized to the complementary strand, the template efficiency of the latter increases several-fold. Under properly selected conditions, highly efficient reverse transcription of the all-ribonucleotide template-primers poly(A).oligo(U), poly(C).oligo(I), and poly(I).oligo(C) can be achieved. "f1 RNA," the RNA strand of an f1 DNA.RNA hybrid, can also serve as template for reverse transcription either after "nicking" of the hybrid with DNase, or after separation from the DNA strand and priming by DNase-treated f1 DNA.