Micallef Peter, Santamaría Manuel Luna, Escobar Mandy, Andersson Daniel, Österlund Tobias, Mouhanna Pia, Filges Stefan, Johansson Gustav, Fagman Henrik, Vannas Christoffer, Ståhlberg Anders
Department of Laboratory Medicine, Institute of Biomedicine, Sahlgrenska Center for Cancer Research, Sahlgrenska Academy, University of Gothenburg, Gothenburg, 413 90, Sweden.
Department of Clinical Genetics and Genomics, Sahlgrenska University Hospital, Gothenburg, 413 45, Sweden.
Genome Biol. 2025 Feb 25;26(1):37. doi: 10.1186/s13059-025-03504-x.
Digital sequencing uses unique molecular identifiers (UMIs) to correct for polymerase induced errors and amplification biases. Here, we design 19 different structured UMIs to minimize the capacity of primers to form non-specific PCR products during library construction using SiMSen-Seq, a PCR-based digital sequencing approach with flexible multiplexing capabilities suitable for tumor-informed mutation analysis. All structured UMI designs demonstrate enhanced assay performance compared with an unstructured reference UMI. The best performing structured UMI design shows significant improvements in all tested aspects of assay and sequencing performance with the ability to reliable detect low variant allele frequencies.
数字测序使用独特分子标识符（UMIs）来校正聚合酶诱导的错误和扩增偏差。在此，我们设计了19种不同结构的UMIs，以在使用SiMSen-Seq进行文库构建期间将引物形成非特异性PCR产物的能力降至最低，SiMSen-Seq是一种基于PCR的数字测序方法，具有适用于肿瘤信息突变分析的灵活多重分析能力。与无结构的参考UMI相比，所有结构化UMI设计均显示出增强的检测性能。性能最佳的结构化UMI设计在检测和测序性能的所有测试方面均有显著改善，能够可靠地检测低变异等位基因频率。
