大肠杆菌素E1质粒DNA在细胞提取物中的复制
Replication of colicin E1 plasmid DNA in cell extracts.
作者信息
Sakakibara Y, Tomizawa J I
出版信息
Proc Natl Acad Sci U S A. 1974 Mar;71(3):802-6. doi: 10.1073/pnas.71.3.802.
Abstract
Cell extracts were prepared from Escherichia coli carrying colicin E1 plasmid. The DNA in extracts was almost exclusively closed-circular DNA of the plasmid. Labeled deoxyribonucleotides were incorporated into DNA in extracts. DNA of colicin E1 plasmid was the sole DNA product, and was composed of completely replicated molecules and a class of replicative intermediates. The intermediates carried an average of approximately two pieces of DNA fragments that had a sedimentation coefficient of approximately 6 S and were not covalently attached to the parental DNA strands. The replication was initiated on closed-circular molecules and the complete molecules were synthesized semiconservatively. The DNA synthesis depended on the four ribonucleoside triphosphates and was sensitive to rifampicin. A round of DNA replication, once initiated, was completed in the presence of rifampicin, indicating that RNA synthesis is involved in the initiation of replication of the plasmid DNA. Most of the replicated molecules were isolated in super-coiled structures. These results indicate that this soluble system is capable of carrying out a complete round of replication of colicin E1 plasmid DNA.
摘要
从携带大肠杆菌素E1质粒的大肠杆菌中制备细胞提取物。提取物中的DNA几乎全是该质粒的闭环DNA。标记的脱氧核糖核苷酸被掺入提取物中的DNA。大肠杆菌素E1质粒的DNA是唯一的DNA产物，由完全复制的分子和一类复制中间体组成。这些中间体平均携带约两条沉降系数约为6S的DNA片段，且这些片段不与亲本DNA链共价连接。复制从闭环分子开始，完整分子通过半保留方式合成。DNA合成依赖于四种核糖核苷三磷酸，且对利福平敏感。一轮DNA复制一旦启动，在利福平存在的情况下也能完成，这表明RNA合成参与了质粒DNA复制的起始。大多数复制的分子以超螺旋结构分离出来。这些结果表明，该可溶性系统能够完成一轮完整的大肠杆菌素E1质粒DNA复制。
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