Demultiplexing and barcode-specific adaptive sampling for nanopore direct RNA sequencing
Abstract Nanopore direct RNA sequencing (dRNA-seq) enables unique insights into RNA biology. However, applications are currently limited by the lack of accurate and cost-effective sample multiplexing. Here we introduce WarpDemuX, an ultra-fast and highly accurate adapter-barcoding and demultiplexing...
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| Format: | Article |
| Language: | English |
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Nature Portfolio
2025-04-01
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| Series: | Nature Communications |
| Online Access: | https://doi.org/10.1038/s41467-025-59102-9 |
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| author | Wiep van der Toorn Patrick Bohn Wang Liu-Wei Marco Olguin-Nava Anne-Sophie Gribling-Burrer Redmond P. Smyth Max von Kleist |
| author_facet | Wiep van der Toorn Patrick Bohn Wang Liu-Wei Marco Olguin-Nava Anne-Sophie Gribling-Burrer Redmond P. Smyth Max von Kleist |
| author_sort | Wiep van der Toorn |
| collection | DOAJ |
| description | Abstract Nanopore direct RNA sequencing (dRNA-seq) enables unique insights into RNA biology. However, applications are currently limited by the lack of accurate and cost-effective sample multiplexing. Here we introduce WarpDemuX, an ultra-fast and highly accurate adapter-barcoding and demultiplexing approach for dRNA-seq with SQK-RNA002 and SQK-RNA004 chemistries. WarpDemuX enhances speed and accuracy by fast processing of the raw nanopore signal, use of a light-weight machine-learning algorithm and design of optimized barcode sets. We demonstrate its utility by performing rapid phenotypic profiling of different SARS-CoV-2 viruses through multiplexed sequencing of longitudinal samples on a single flowcell, identifying systematic differences in transcript abundance and poly(A) tail lengths during infection. Additionally, integrating WarpDemuX into sequencing control software enables real-time enrichment of target molecules through barcode-specific adaptive sampling, which we demonstrate by enriching low abundance viral RNA. In summary, WarpDemuX represents a broadly applicable, high-performance, economical multiplexing solution for dRNA-seq, facilitating advanced (epi-) transcriptomic research. |
| format | Article |
| id | doaj-art-8f609053842d40d1abd91a8f60c4b2f4 |
| institution | Kabale University |
| issn | 2041-1723 |
| language | English |
| publishDate | 2025-04-01 |
| publisher | Nature Portfolio |
| record_format | Article |
| series | Nature Communications |
| spelling | doaj-art-8f609053842d40d1abd91a8f60c4b2f42025-08-20T03:53:32ZengNature PortfolioNature Communications2041-17232025-04-0116111310.1038/s41467-025-59102-9Demultiplexing and barcode-specific adaptive sampling for nanopore direct RNA sequencingWiep van der Toorn0Patrick Bohn1Wang Liu-Wei2Marco Olguin-Nava3Anne-Sophie Gribling-Burrer4Redmond P. Smyth5Max von Kleist6Systems Medicine of Infectious Disease (P5), Robert Koch InstituteHelmholtz Institute for RNA-based Infection Research, Helmholtz Centre for Infection ResearchSystems Medicine of Infectious Disease (P5), Robert Koch InstituteHelmholtz Institute for RNA-based Infection Research, Helmholtz Centre for Infection ResearchHelmholtz Institute for RNA-based Infection Research, Helmholtz Centre for Infection ResearchHelmholtz Institute for RNA-based Infection Research, Helmholtz Centre for Infection ResearchSystems Medicine of Infectious Disease (P5), Robert Koch InstituteAbstract Nanopore direct RNA sequencing (dRNA-seq) enables unique insights into RNA biology. However, applications are currently limited by the lack of accurate and cost-effective sample multiplexing. Here we introduce WarpDemuX, an ultra-fast and highly accurate adapter-barcoding and demultiplexing approach for dRNA-seq with SQK-RNA002 and SQK-RNA004 chemistries. WarpDemuX enhances speed and accuracy by fast processing of the raw nanopore signal, use of a light-weight machine-learning algorithm and design of optimized barcode sets. We demonstrate its utility by performing rapid phenotypic profiling of different SARS-CoV-2 viruses through multiplexed sequencing of longitudinal samples on a single flowcell, identifying systematic differences in transcript abundance and poly(A) tail lengths during infection. Additionally, integrating WarpDemuX into sequencing control software enables real-time enrichment of target molecules through barcode-specific adaptive sampling, which we demonstrate by enriching low abundance viral RNA. In summary, WarpDemuX represents a broadly applicable, high-performance, economical multiplexing solution for dRNA-seq, facilitating advanced (epi-) transcriptomic research.https://doi.org/10.1038/s41467-025-59102-9 |
| spellingShingle | Wiep van der Toorn Patrick Bohn Wang Liu-Wei Marco Olguin-Nava Anne-Sophie Gribling-Burrer Redmond P. Smyth Max von Kleist Demultiplexing and barcode-specific adaptive sampling for nanopore direct RNA sequencing Nature Communications |
| title | Demultiplexing and barcode-specific adaptive sampling for nanopore direct RNA sequencing |
| title_full | Demultiplexing and barcode-specific adaptive sampling for nanopore direct RNA sequencing |
| title_fullStr | Demultiplexing and barcode-specific adaptive sampling for nanopore direct RNA sequencing |
| title_full_unstemmed | Demultiplexing and barcode-specific adaptive sampling for nanopore direct RNA sequencing |
| title_short | Demultiplexing and barcode-specific adaptive sampling for nanopore direct RNA sequencing |
| title_sort | demultiplexing and barcode specific adaptive sampling for nanopore direct rna sequencing |
| url | https://doi.org/10.1038/s41467-025-59102-9 |
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