Velappan Nileena, Sblattero Daniele, Chasteen Leslie, Pavlik Peter, Bradbury Andrew R M
Biosciences Division, Los Alamos National Laboratory, Los Alamos, NM 87545, USA.
Protein Eng Des Sel. 2007 Jul;20(7):309-13. doi: 10.1093/protein/gzm005. Epub 2007 Mar 1.
It is generally accepted that plasmids containing the same origin of replication are incompatible. We have re-examined this concept in terms of the plasmid copy number, by introducing plasmids containing the same origin of replication and different antibiotic resistance genes into bacteria. By selecting for resistance to only one antibiotic, we were able to examine the persistence of plasmids carrying resistances to other antibiotics. We find that plasmids are not rapidly lost, but are able to persist in bacteria for multiple overnight growth cycles, with some dependence upon the nature of the antibiotic selected for. By carrying out the experiments with different origins of replication, we have been able to show that higher copy number leads to longer persistence, but even with low copy plasmids, persistence occurs to a significant degree. This observation holds significance for the field of protein engineering, as the presence of two or more plasmids within bacteria weakens, and confuses, the connection between screened phenotype and genotype, with the potential to wrongly assign specific phenotypes to incorrect genotypes.
一般认为，含有相同复制起点的质粒是不相容的。我们通过将含有相同复制起点和不同抗生素抗性基因的质粒导入细菌，从质粒拷贝数的角度重新审视了这一概念。通过仅选择对一种抗生素的抗性，我们能够检测携带其他抗生素抗性的质粒的持久性。我们发现质粒不会迅速丢失，而是能够在细菌中持续多个过夜生长周期，这在一定程度上取决于所选择的抗生素的性质。通过使用不同的复制起点进行实验，我们能够表明较高的拷贝数会导致更长的持久性，但即使是低拷贝质粒，也会有相当程度的持久性。这一观察结果对蛋白质工程领域具有重要意义，因为细菌中存在两种或更多种质粒会削弱并混淆筛选出的表型与基因型之间的联系，有可能将特定表型错误地归因于错误的基因型。
