The chromosome of bacteriophage T5. II. Arrangement of the single-stranded DNA fragments in the T5⁺ and T5st(O) chromosomes

Denatured bacteriophage T5 DNA contains a large number of single-stranded DNA fragments which have been separated by agarose gel electrophoresis and classified as "major" or "minor" species on the basis of their relative abundances (Hayward & Smith, 1972). For further study of these fragments we have centrifuged denatured T5 DNA in CsCl density-gradients in the presence of poly(G). Gel electrophoretic analysis of fractions from these gradients shows that the 37.0 and 13.9 million major fragments of T5⁺ DNA and the 35.3 and 17.2 million of T5st(O) DNA are found in the high buoyant density regions. The other fragments vary in the extent of their interactions with poly(G) and a minor fragment, which has anomalous electrophoretic properties, exhibits the strongest poly(G) interaction. Agarose gel electrophoresis has also been used for analysing the results of hybridization experiments with isolated major fragments and for studying the distribution of single-strand fragments in the double-strand breakage products of sheared T5 DNA. From the results of these experiments we have proposed a model for the arrangment of major single-strand fragments in the duplex DNA molecules of T5⁺ and T5st(O). In our model, both molecules contain one intact strand, and 3.8 and 14.5 million fragments at opposite ends of the other strand, but T5⁺ contains three "primary" interruptions and T5st(O) contains only two. This difference is explained by a deletion in T5st(O) DNA which eliminates the central T5⁺ interruption and creates the new 17.2 million fragment of T5st(O) from the 5.1 and 13.9 million fragments of T5⁺. The first break by shearing in duplex T5 DNA occurs in the intact strand at a position approximately opposite the interruption separating the 14.5 million fragment from the rest of the "fragmented" strand. We have also studied the single strands of first-step-transfer DNA and suggest that the division between first-step-transfer DNA and the rest of the molecule is not associated with a single-strand interruption.