Oxford Nanopore Technologies (ONT) sequencing platforms currently offer two approaches to whole-genome native-DNA library preparation: ligation and rapid. In this study, we compared these two approaches for bacterial whole-genome sequencing, with a specific aim of assessing their ability to recover small plasmid sequences. To do so, we sequenced DNA from seven plasmid-rich bacterial isolates in three different ways: ONT ligation, ONT rapid and Illumina. Using the Illumina read depths to approximate true plasmid abundance, we found that small plasmids (<20 kbp) were underrepresented in ONT ligation read sets (by a mean factor of ~4) but were not underrepresented in ONT rapid read sets. This effect correlated with plasmid size, with the smallest plasmids being the most underrepresented in ONT ligation read sets. We also found lower rates of chimaeric reads in the rapid read sets relative to ligation read sets. These results show that when small plasmid recovery is important, ONT rapid library preparations are preferable to ligation-based protocols.

中文翻译：

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通过牛津纳米孔测序恢复小质粒序列

Oxford Nanopore Technologies (ONT) 测序平台目前提供两种全基因组天然 DNA 文库制备方法：连接和快速。在本研究中，我们比较了这两种细菌全基因组测序的方法，具体目的是评估它们恢复小质粒序列的能力。为此，我们以三种不同的方式对来自七种富含质粒的细菌分离株的 DNA 进行了测序：ONT 结扎、ONT rapid 和 Illumina。使用 Illumina 读取深度来近似真实的质粒丰度，我们发现小质粒（<20 kbp）在 ONT 连接读取集中的代表性不足（平均系数约为 4），但在 ONT 快速读取集中并没有被低估。这种效应与质粒大小相关，最小的质粒在 ONT 连接读取集中的代表性最小。我们还发现，与连接读取集相比，快速读取集中的嵌合读取率较低。这些结果表明，当小质粒回收很重要时，ONT 快速文库制备优于基于连接的方案。