Harmer Christopher J, Hall Ruth M
School of Life and Environmental Sciences, The University of Sydney, Sydney, New South Wales, Australia.
Mol Microbiol. 2017 Nov;106(3):409-418. doi: 10.1111/mmi.13774. Epub 2017 Sep 4.
We recently proposed a model for targeted, conservative cointegrate formation between DNA molecules each containing a copy of IS26, that involves Tnp26-catalyzed strand exchange occurring at either the two left ends or the two right ends of the IS. Here, this model was validated by altering the bases at the outer left terminus, right terminus or both termini of one IS26. The correct bases at both ends were required in the untargeted replicative mode. However, when only one end was altered in one participating IS the frequency of targeted, conservative cointegrate formation was not reduced. The distribution of the altered bases in the cointegrates confirmed that the reaction occurred at the end where the terminal bases of both IS were correct, and cointegrates were not formed when both ends of the same IS were altered. The terminal bases of the active IS26 were also required to support deletion of the aphA1a translocatable unit (TU) from Tn4352B. The choices made by an incoming TU with a wild-type IS26 when the target plasmid included one wild-type IS26 and one with a frameshift in tnp26 demonstrated that Tnp26 exhibits a strong preference for cis action.
我们最近提出了一个模型，用于在每个都含有一份IS26拷贝的DNA分子之间进行靶向、保守的共整合体形成，该模型涉及由Tnp26催化的链交换，其发生在IS的两个左端或两个右端。在此，通过改变一个IS26的外左端、右端或两端的碱基，对该模型进行了验证。在非靶向复制模式下，两端都需要正确的碱基。然而，当参与的一个IS中只有一端被改变时，靶向、保守的共整合体形成频率并未降低。共整合体中改变碱基的分布证实，反应发生在两个IS的末端碱基都正确的一端，并且当同一个IS的两端都被改变时，不会形成共整合体。活性IS26的末端碱基对于支持从Tn4352B中删除aphA1a易位单元（TU）也是必需的。当目标质粒包含一个野生型IS26和一个在tnp26中发生移码的IS26时，带有野生型IS26的传入TU所做出的选择表明，Tnp26表现出强烈的顺式作用偏好。
