van der Toorn Wiep, Naarmann-de Vries Isabel S, Liu-Wei Wang, Dieterich Christoph, von Kleist Max
Systems Medicine of Infectious Disease (P5), Robert Koch Institute, 13353 Berlin, Germany.
Department of Mathematics and Computer Science, Freie Universität Berlin, 14195 Berlin, Germany.
Nucleic Acids Res. 2025 Sep 5;53(17). doi: 10.1093/nar/gkaf873.
Transfer RNA (tRNA) plays an essential role in protein translation, and tRNA modifications are important to their function. Recently, nanopore direct RNA sequencing (dRNA-seq) has shown promising results in the detection of complex tRNA modifications. However, its wider adoption in the tRNA field has been limited by a lack of (de)multiplexing solutions. Here, we present WarpDemuX-tRNA: an extension to the WarpDemuX method specifically optimized for multiplexed nanopore tRNA sequencing. Using consensus-based signal analysis using (soft) dynamic time warping and barycenter averaging, our approach improves barcode feature generation and achieves more robust barcode identification. WarpDemuX-tRNA outperforms the original method and achieves 99% precision and 95% recovery for four barcodes, while reducing computational complexity and runtime to 6 min per one million reads. WarpDemuX-tRNA is an open-source and free-to-use solution to high-throughput nanopore tRNA sequencing, facilitating more accessible, cost-effective, and high-throughput studies of tRNA modifications and their regulatory mechanisms.
转运RNA（tRNA）在蛋白质翻译中起着至关重要的作用，tRNA修饰对其功能很重要。最近，纳米孔直接RNA测序（dRNA-seq）在检测复杂的tRNA修饰方面显示出了有前景的结果。然而，由于缺乏（去）多路复用解决方案，它在tRNA领域的更广泛应用受到了限制。在这里，我们介绍WarpDemuX-tRNA：WarpDemuX方法的一个扩展，专门针对多路复用纳米孔tRNA测序进行了优化。使用基于一致性的信号分析，采用（软）动态时间规整和重心平均，我们的方法改进了条形码特征生成，并实现了更稳健的条形码识别。WarpDemuX-tRNA优于原始方法，对于四个条形码实现了99%的精度和95%的回收率，同时将计算复杂度和运行时间降低到每百万读数6分钟。WarpDemuX-tRNA是一种开源且免费使用的高通量纳米孔tRNA测序解决方案，有助于更便捷、经济高效且高通量地研究tRNA修饰及其调控机制。
