The nucleotide sequence at the 3′-termini of three major T5 DNA fragments

Abstract

Abstract T5 DNA contains several site-specific nicks in one of the two strands. Since the 5′-nucleotide at the majority of the nicks is dGMP, the 3′-nucleotide can be labeled using Escherichia coli DNA polymerase I and [α- 32 P]dGTP. Terminally labeled single-stranded fragments were separated by agarose-gel electrophoresis of denatured DNA. A 40- to 45-nucleotide sequence at the 3′-termini of three of the major single-stranded fragments was deduced from a combination of two-dimensional homochromatography and the Maxam-Gilbert gel-sequencing technique. The sequence immediately adjacent to the nicks is asymmetric, but two overlapping regions of twofold rotational symmetry occur with axes that are 11–15 nucleotides to the 3′-side of each nick. Other regions of high GC or AT content are conserved among the three sequences. The possible relevance of these sequences with regard to the putative site-specific endonucleases of T5 and temporal regulation of T5 mRNA synthesis is discussed.