Shlomai J, Strauss B, Asher Y, Friedmann A, Becker Y
J Gen Virol. 1976 Aug;32(2):189-204. doi: 10.1099/0022-1317-32-2-189.
The nature of the DNA molecules synthesized in nuclei of herpes simplex virus (HSV)-infected cells in vivo and in vitro was studied by chromatography on BND-cellulose columns after shearing to DNA fragments of 10 to 20 X 10(6) daltons. The incorporation of labelled precursors occurs in the DNA fragments containing single-stranded regions, presumably the replication forks. Prolongation of DNA synthesis leads to the accumulation of labelled DNA fragments that lack single-stranded sequences. Analysis of the isolated DNA fragments by density centrifugation in CSCl gradients revealed that most of the labelled DNA molecules are of virus specificity and the minority are cellular DNA fragments. Double-stranded virus DNA fragments and virus DNA fragments containing single-stranded sequences band in CSCl gradients at a density of 1-718 g/ml, the density of virion DNA. This suggests that the replicating HSV DNA molecules have the same density as the virion DNA and contain relatively little single-stranded DNA. The synthesis of HSV DNA molecules under in vitro conditions in isolated nuclei occurs by incorporation of the precursors into DNA fragments with single-stranded regions. The synthesis of cellular DNA in nuclei from hydroxyurea and cytosine arabinoside treated cells also occurs by elongation of nascent DNA chains.
通过在BND-纤维素柱上进行色谱分析，研究了单纯疱疹病毒（HSV）感染细胞在体内和体外细胞核中合成的DNA分子的性质，这些DNA分子被剪切成10至20×10⁶道尔顿的DNA片段。标记前体的掺入发生在含有单链区域的DNA片段中，推测这些单链区域为复制叉。DNA合成的延长导致缺乏单链序列的标记DNA片段的积累。通过在CSCl梯度中进行密度离心对分离的DNA片段进行分析，结果显示大多数标记的DNA分子具有病毒特异性，少数为细胞DNA片段。双链病毒DNA片段和含有单链序列的病毒DNA片段在CSCl梯度中的密度为1.718 g/ml，即病毒粒子DNA的密度。这表明正在复制的HSV DNA分子与病毒粒子DNA具有相同的密度，并且含有相对较少的单链DNA。在体外条件下，分离细胞核中HSV DNA分子的合成是通过将前体掺入到具有单链区域的DNA片段中实现的。羟基脲和阿糖胞苷处理的细胞的细胞核中细胞DNA的合成也是通过新生DNA链的延伸来进行的。
