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The Synthesis and Substructure of Herpesvirus DNA: the Distribution of Alkali-labile Single Strand Interruptions in HSV-1 DNA

Institute of Virology, University of Glasgow, Glasgow, Scotland

Denatured DNA of herpes simplex virus was released from the particles using an alkaline detergent, Decon-75. The largest single strands sedimented on alkaline sucrose gradients with a mol. wt. of 47.2 ± 0.33 x 10⁶, slightly less than half the value calculated for the intact duplex (104 x 10⁶). About 50% of the DNA was found in fragments which sedimented slower than this in a heterogeneous manner. On agarose gel electrophoresis the largest strands migrated with a mol. wt. of 40 x 10⁶. The reason for this difference is not known but since the individual strands of T 4 DNA were shown to migrate with slightly different R_r values, factors other than mol. wt. may affect the migration of single-stranded DNA. Two fragments of mol. wt. 35 x 10⁶ and 30 x 10⁶ were observed but the rest of the fragments remained unresolved by this technique. Virus DNA associated with the nucleus of infected cells had a much lower mol. wt. than particle DNA (3.2 x 10⁶). Although this value increased if the DNA was isolated from intact cells, the average sedimentation coefficient of nuclear virus DNA was never as high as the largest strands of particle DNA.

When the largest single strands from particles were prepared by sucrose gradient fractionation they exhibited a unimodal mol. wt. distribution after both sedimentation and electrophoretic analysis. These ‘intact’ single strands were annealed and analysed by banding in CsCl gradients and by analyses employing the Neurospora crassa endonuclease. The results show that the ‘intact’ strands reassociated with the same kinetics and to the same extent as total virus DNA, suggesting that both strands of the duplex were present in equal amounts.

Received 4 May 1973; accepted 23 July 1973.

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