Regions of broad-host-range plasmid RK2 which are essential for replication and maintenance
Regions of broad-host-range plasmid RK2 which are essential for replication and maintenance
复制标题
广泛宿主范围质粒 RK2 的复制和维持所必需的区域
DOI：
10.1128/jb.141.1.213-222.1980
复制
发表时间：
1980
影响因子：
3.2
通讯作者：
D. Helinski
中科院分区：
文献类型：
--
作者：
Christopher M Thomas;R. Meyer;D. Helinski
广泛的主机范围的质粒RK2（56个酶）的地图上的裂解位点进行了测定的BglII，PstI，和SmaI限制性内切酶，和四环素和氨苄青霉素抗性的决定因素进行了本地化。切割位点聚集在耐药基因处或附近。本地化的质粒复制和维护在大肠杆菌中所需的区域，我们删除了非必需的区域RK2的部分消化与限制性内切酶HaeII产生小的衍生物。获得的最小的稳定复制子含有5个RK2的HaeII片段，总共5.4个酶。这些片段源自RK 2的三个区域，这些区域被抗生素耐药基因彼此分开。这些HaeII片段之一（0.75内切酶）具有复制起点的预期特性。发现位于RK2的两个独立区域中的外部四个片段提供反式功能，其允许携带复制起点的HaeII片段的复制。这些结果表明，至少有两个质粒编码的基因，能够在反式，和一个复制起点所需的RK2复制和维护。
The sites of cleavage on the map of the broad-host-range plasmid RK2 (56 kilobases) were determined for the BglII, PstI, and SmaI restriction enzymes, and the determinants for tetracycline and ampicillin resistance were localized. The cleavage sites were clustered at or near the drug resistance genes. To localize regions required for plasmid replication and maintenance in Escherichia coli, we deleted nonessential regions of RK2 by partial digestion with the restriction endonuclease HaeII to produce small derivatives. The smallest stable replicon obtained contained five HaeII fragments of RK2 which total 5.4 kilobases. These fragments were derived from three regions of RK2 that are separated from each other by antibiotic resistance genes. One of these HaeII fragments (0.75 kilobases) has the properties expected of the origin of replication. The outer four fragments, located in two separate regions of RK2, were found to provide, in trans, functions that permit the replication of the HaeII fragment carrying the origin of the replication. These results indicate that at least two plasmid-encoded genes, capable of acting in trans, and a replication origin are required for RK2 replication and maintenance.