Gourlie B B, Krauss M R, Buckler-White A J, Benbow R M, Pigiet V
J Virol. 1981 Jun;38(3):805-14. doi: 10.1128/JVI.38.3.805-814.1981.
Polyoma virus minichromosomes were isolated from infected 3T6 cells by hypotonic extraction of isolated nuclei. The kinetics of in vitro DNA synthesis in the nuclear extract was similar to that observed with intact nuclei. The majority of the products of in vitro DNA synthesis sedimented with replicative intermediate (RI) minichromosomes and migrated as two bands (RI-a and RI-b) on 1.4% agarose gels. The kinetics of deoxynucleotide monophosphate incorporation into these species was consistent with the existence of several rate-limiting steps in in vitro replication by polyoma minichromosomes. Electron microscope analysis showed that the RI-a band consisted almost entirely of RI theta structures ranging from 46 to 87% replicated, with one-half of all theta structures 67 +/- 4% replicated. The RI-b material was more complex, consisting of sigma and alpha structures with tails ranging from 7 to 114% of polyoma genome length and, less frequently, of linked and multiple linked dimeric structures.
通过低渗提取分离的细胞核，从感染的3T6细胞中分离出多瘤病毒微型染色体。核提取物中体外DNA合成的动力学与完整细胞核中观察到的相似。体外DNA合成的大多数产物与复制中间体（RI）微型染色体一起沉淀，并在1.4%琼脂糖凝胶上迁移为两条带（RI-a和RI-b）。脱氧核苷酸单磷酸掺入这些物种的动力学与多瘤微型染色体体外复制中存在几个限速步骤一致。电子显微镜分析表明，RI-a带几乎完全由复制率为46%至87%的RIθ结构组成，所有θ结构的一半复制率为67±4%。RI-b物质更复杂，由σ和α结构组成，尾巴长度为多瘤病毒基因组长度的7%至114%，较少见的是连接和多重连接的二聚体结构。
