School of Life Sciences, Medical School, Queens Medical Centre, University of Nottingham, Nottingham NG72RD, United Kingdom.
Bioinformatics. 2024 Mar 29;40(4). doi: 10.1093/bioinformatics/btae141.
Oxford Nanopore Technologies (ONT) sequencers enable real-time generation of sequence data, which allows for concurrent analysis during a run. Adaptive sampling leverages this real-time capability in extremis, rejecting or accepting reads for sequencing based on assessment of the sequence from the start of each read. This functionality is provided by ONT's software, MinKNOW (Oxford Nanopore Technologies). Designing and developing software to take advantage of adaptive sampling can be costly in terms of sequencing consumables, using precious samples and preparing sequencing libraries. MinKNOW addresses this in part by allowing the replay of previously sequenced runs for testing. However, as we show, the sequencing output only partially changes in response to adaptive sampling instructions. Here we present Icarust, a tool enabling more accurate approximations of sequencing runs. Icarust recreates all the required endpoints of MinKNOW to perform adaptive sampling and writes output compatible with current base-callers and analysis pipelines. Icarust serves nanopore signal simulating a MinION or PromethION flow cell experiment from any reference genome using either R9 or R10 pore models. We show that simulating sequencing runs with Icarust provides a realistic testing and development environment for software exploiting the real-time nature of Nanopore sequencing.
All code is open source and freely available here-https://github.com/LooseLab/Icarust. Icarust is implemented in Rust, with a docker container also available. The data underlying this article will be shared on reasonable request to the corresponding author.
牛津纳米孔技术(ONT)测序仪能够实时生成序列数据,这使得在运行过程中可以进行并发分析。自适应采样利用了这种实时能力,根据每个读取起始时的序列评估,拒绝或接受读取进行测序。此功能由 ONT 的软件 MinKNOW(牛津纳米孔技术)提供。为了利用自适应采样设计和开发软件可能会在测序耗材、使用珍贵样本和准备测序文库方面造成巨大的成本。MinKNOW 通过允许重播以前测序的运行来部分解决此问题。但是,正如我们所展示的,测序输出仅部分响应自适应采样指令而改变。在这里,我们介绍了 Icarust,这是一种能够实现更准确的测序运行模拟的工具。Icarust 重新创建了 MinKNOW 执行自适应采样所需的所有端点,并编写与当前碱基调用器和分析管道兼容的输出。Icarust 使用 R9 或 R10 孔模型,从任何参考基因组模拟 MinION 或 PromethION 流控芯片实验的纳米孔信号。我们表明,使用 Icarust 模拟测序运行可以为利用 Nanopore 测序实时性的软件提供现实的测试和开发环境。
所有代码都是开源的,并可在此处免费获得-https://github.com/LooseLab/Icarust。Icarust 是用 Rust 实现的,同时也提供了一个 Docker 容器。本文所依据的数据将根据相应作者的合理请求共享。