TIRE-seq simplifies transcriptomics via integrated RNA capture and library preparation
RNA sequencing (RNA-seq) is widely used in biomedical research, advancing our understanding of gene expression across biological systems. Traditional methods require upstream RNA extraction from biological inputs, adding time and expense to workflows. We developed TIRE-seq (Turbocapture Integrated RNA Expression Sequencing) to address these challenges. TIRE-seq integrates mRNA purification directly into library preparation, eliminating the need for a separate extraction step. This streamlined approach reduces turnaround time, minimizes sample loss, and improves data quality. A comparative study with the widely used Prime-seq protocol demonstrates TIRE-seq's superior sequencing efficiency with crude cell lysates as inputs. TIRE-seq's utility was demonstrated across three biological applications. It captured transcriptional changes in stimulated human T cells, revealing activation-associated gene expression profiles. It also identified key genes driving murine dendritic cell differentiation, providing insights into lineage commitment. Lastly, TIRE-seq analyzed the dose-response and time-course effects of temozolomide on patient-derived neurospheres, identifying differentially expressed genes and enriched pathways linked to the drug's mechanism of action. With its simplified workflow and high sequencing efficiency, TIRE-seq offers a cost-effective solution for large-scale gene expression studies across diverse biological systems.
Authors
- Patrick O’Keeffe (ORCID: https://orcid.org/0000-0002-8676-4436)
- Daniela Amann‐Zalcenstein (ORCID: https://orcid.org/0000-0003-4928-1846)
- Matthew E. Ritchie (ORCID: https://orcid.org/0000-0002-7383-0609)
- Hui Shi Saw
- Ryan Cross (ORCID: https://orcid.org/0000-0002-8573-8551)
- Tracey M. Baldwin (ORCID: https://orcid.org/0000-0002-8919-5822)
- Daniel V. Brown (ORCID: https://orcid.org/0000-0002-3594-5851)
- David Squire (ORCID: https://orcid.org/0000-0001-6738-8271)
- Cindy Audiger (ORCID: https://orcid.org/0000-0001-9087-9036)
- Sam W.Z. Olechnowicz (ORCID: https://orcid.org/0000-0001-7879-7708)
- James Richard Whittle (ORCID: https://orcid.org/0000-0002-4740-2037)
- Stephen J Leslie (ORCID: https://orcid.org/0000-0002-1403-4733)
- Misty R. Jenkins (ORCID: https://orcid.org/0000-0001-6564-2715)
- Shalin H. Naik (ORCID: https://orcid.org/0000-0003-0299-3301)
- Rory Bowden (ORCID: https://orcid.org/0000-0001-8596-0366)
- Sarah A. Best (ORCID: https://orcid.org/0000-0002-4232-9432)
- Jafar S. Jabbari (ORCID: https://orcid.org/0000-0002-6302-4857)
- Zachery Moore (ORCID: https://orcid.org/0000-0002-0898-0483)
- Peter F. Hickey (ORCID: https://orcid.org/0000-0002-8153-6258)
- Yupei You (ORCID: https://orcid.org/0000-0001-7302-3033)
- Changqing Wang (ORCID: https://orcid.org/0000-0001-7868-3882)
- Yasmin Nouri
- Saskia Freytag (ORCID: https://orcid.org/0000-0002-2185-7068)
Institutions
- The University of Melbourne (AU)
Publication Details
- Journal
- Scientific Reports
- Published
- 2025-05-02
- DOI
- https://doi.org/10.1038/s41598-025-98282-8
- Citations
- 1
- Primary Topic
- RNA modifications and cancer
- Type
- article
- Field-Weighted Citation Impact
- 0.81