Meijer W J, van der Lelie D, Venema G, Bron S
Department of Genetics, Centre for Biological Sciences, Kerklaan, Haren, The Netherlands.
Plasmid. 1995 Mar;33(2):91-9. doi: 10.1006/plas.1995.1011.
The effects of the single-strand origins (SSOs) of the broad-host-range streptococcal plasmid pMV158 on (i) the conversion of its single-stranded (ss) DNA replication intermediates to double-stranded (ds) plasmid DNA and (ii) its maintenance were analyzed. pMV158 is distinguished from most other plasmids that replicate by the rolling-circle mechanism by the presence of two single-strand origins of replication, palA and palU. In this paper the results obtained with Lactococcus lactis are presented; complementary studies with Bacillus subtilis are presented in the accompanying paper (Meijer et al., 1995). In the presence of both SSOs, no ss plasmid DNA was observed in L. lactis. The removal of either palA or palU resulted in the appearance of low amounts of ssDNA. High amounts of ssDNA were detected, however, when both SSOs were deleted. The results indicated that both SSOs were active, albeit that palU was the most effective of the two. In the presence of both SSOs, the plasmid was stably maintained in L. lactis under nonselective growth conditions. Also, the derivatives containing only one of the two SSOs were maintained rather stably. In contrast, the derivative devoid of both SSOs was poorly maintained. It was concluded that, in the absence of a functional SSO, the generation of large amounts of ssDNA drastically reduces the maintenance of pMV158 in L. lactis. The results also showed that the presence of the plasmid-located mob gene, required for conjugative mobilization, was involved neither in the accumulation of ssDNA nor in the maintenance of pMV158.
分析了广宿主范围的链球菌质粒pMV158的单链复制起点（SSO）对（i）其单链（ss）DNA复制中间体转化为双链（ds）质粒DNA的影响以及（ii）对其稳定性的影响。pMV158与大多数通过滚环机制复制的其他质粒不同，它有两个单链复制起点palA和palU。本文展示了在乳酸乳球菌中获得的结果；在随附论文（Meijer等人，1995年）中展示了与枯草芽孢杆菌的互补研究。在两个SSO都存在的情况下，在乳酸乳球菌中未观察到单链质粒DNA。去除palA或palU中的任何一个都会导致少量单链DNA的出现。然而，当两个SSO都缺失时，检测到大量单链DNA。结果表明两个SSO都是有活性的，尽管palU在两者中是最有效的。在两个SSO都存在的情况下，该质粒在非选择性生长条件下能在乳酸乳球菌中稳定维持。此外，仅含有两个SSO之一的衍生物也能相当稳定地维持。相比之下，不含两个SSO的衍生物维持效果较差。得出的结论是，在没有功能性SSO的情况下，大量单链DNA的产生会大幅降低pMV158在乳酸乳球菌中的稳定性。结果还表明，参与接合转移所需的位于质粒上的mob基因，既不参与单链DNA的积累，也不参与pMV158的稳定性维持。
