Troester H, Bub S, Hunziker A, Trendelenburg M F
German Cancer Research Center, aBiomedical Structure Analysis (A0600), Im Neuenheimer Feld 280, 69120, Heidelberg, Germany.
Gene. 2000 Nov 27;258(1-2):95-108. doi: 10.1016/s0378-1119(00)00420-0.
In this study we report on the stabilization of short direct repetitive DNA elements. We arranged a 20 bp SK-primer element in a direct repeat manner within the cloning vector pBluescript KS (+). This resulted in an array of 27 direct repeats consisting of 24 bp units. We show that this plasmid could only be propagated without deletion of repeats in dam mutant Escherichia coli hosts, whereas all efforts to use strains that were defective in the methylation-dependent restriction system and the recA- or the mismatch repair-dependent deletion system failed. The deletions always affected whole repeat units and not parts of them, leading to an unpredictable reduction of the unit number down to a range of between 12 and two during propagation. We conclude that a Dam methylation-dependent, but recA- and mismatch repair-independent, deletion mechanism caused the DNA rearrangements without an obvious involvement of the known methylated-DNA restriction systems.
在本研究中，我们报道了短直接重复DNA元件的稳定性。我们将一个20 bp的SK引物元件以直接重复的方式排列在克隆载体pBluescript KS(+)内。这产生了一个由24 bp单位组成的27个直接重复序列的阵列。我们发现，这种质粒只能在dam突变型大肠杆菌宿主中进行无重复序列缺失的扩增，而使用在甲基化依赖的限制系统以及recA或错配修复依赖的缺失系统中存在缺陷的菌株的所有尝试均告失败。缺失总是影响整个重复单位而非其部分，导致在扩增过程中单位数量不可预测地减少至12到2个的范围。我们得出结论，一种依赖Dam甲基化但不依赖recA和错配修复的缺失机制导致了DNA重排，而已知的甲基化DNA限制系统未明显参与其中。
