Volovitch M, Drugeon C, Yot P
Nucleic Acids Res. 1978 Aug;5(8):2913-25. doi: 10.1093/nar/5.8.2913.
The Cauliflower Mosaic Virus (CaMV) genome is a double-stranded DNA molecule of about 5 million daltons. Native DNA molecules appear heterogeneous when analysed by gel electrophoresis. We have examined the nature of this apparent heterogeneity. Besides, this genome is shown here to contain three single-stranded breaks, as revealed by different denaturation experiments: heating at 75 degrees C, treatment with NaOH or dimethyl sulfoxide (DMSO). Labelling with terminal transferase proves that the 3' ends at these interruptions all have free hydroxyl groups. Electron microscopy and alkaline gel electrophoresis indicate that these three discontinuities are shared by both strands, and that they are not randomly located. S1 nuclease is active on CaMV DNA and generates three fragments. The comparison between the sizes of these fragments and of the products of denaturation leads us to consider that S1 acts at the level of the interruptions. We have determined that two of them, distant by one third genome unit, are in the same strand; the other is in the opposite strand, distant by one sixth genome unit from the nearest other one. The combined use of restriction enzymes and S1 nuclease has enabled us to locate these three discontinuities on the restriction map of the CaMV genome that we have otherwise established.
花椰菜花叶病毒（CaMV）基因组是一个约500万道尔顿的双链DNA分子。当通过凝胶电泳分析时，天然DNA分子呈现出异质性。我们研究了这种明显异质性的本质。此外，通过不同的变性实验（75℃加热、用氢氧化钠或二甲基亚砜（DMSO）处理）表明，该基因组含有三个单链断裂处。用末端转移酶标记证明这些中断处的3'末端都有游离羟基。电子显微镜和碱性凝胶电泳表明这三个不连续处两条链都有，且它们不是随机定位的。S1核酸酶对CaMV DNA有活性并产生三个片段。这些片段大小与变性产物大小的比较使我们认为S1作用于中断处。我们已确定其中两个相距一个基因组单位的三分之一，位于同一条链上；另一个在相反链上，与最近的一个相距一个基因组单位的六分之一。限制性内切酶和S1核酸酶的联合使用使我们能够在我们另行建立的CaMV基因组限制性图谱上定位这三个不连续处。
