Academic Journal
A novel one-step multiplex PCR method (OS-MPCR) for efficient NGS library preparation in cancer.
| Τίτλος: | A novel one-step multiplex PCR method (OS-MPCR) for efficient NGS library preparation in cancer. |
|---|---|
| Συγγραφείς: | Feng H; Department of Pathology, Zibo First Hospital, Zibo, Shandong, China., Hu X; Department of Pathology, Zibo First Hospital, Zibo, Shandong, China., Zhang L; Department of Pathology, Zibo First Hospital, Zibo, Shandong, China., Zhang Q; Department of Pathology, Zibo First Hospital, Zibo, Shandong, China., Wang S; Shanghai Yijian Health Technology Co., Ltd., Shanghai, China., Lv B; Shanghai Yijian Health Technology Co., Ltd., Shanghai, China., Zhang Y; Shanghai Yijian Health Technology Co., Ltd., Shanghai, China., Shi D; Shanghai Yijian Health Technology Co., Ltd., Shanghai, China., Zhou P; Department of Pathology, Zibo First Hospital, Zibo, Shandong, China. |
| Πηγή: | PloS one [PLoS One] 2026 Sep 15; Vol. 21 (9), pp. e0357776. Date of Electronic Publication: 2026 Sep 15 (Print Publication: 2026). |
| Τύπος έκδοσης: | Journal Article |
| Γλώσσα: | English |
| Στοιχεία περιοδικού: | Publisher: Public Library of Science Country of Publication: United States NLM ID: 101285081 Publication Model: eCollection Cited Medium: Internet ISSN: 1932-6203 (Electronic) Linking ISSN: 19326203 NLM ISO Abbreviation: PLoS One Subsets: MEDLINE |
| Imprint Name(s): | Original Publication: San Francisco, CA : Public Library of Science |
| Ιατρικοί όροι (MeSH): | Multiplex Polymerase Chain Reaction*/methods , High-Throughput Nucleotide Sequencing*/methods , Lung Neoplasms*/genetics , Gene Library*, Humans |
| Περίληψη: | Background: Next-generation sequencing (NGS) is essential for precision oncology, yet its clinical implementation is often hindered by limited sample quantity, complex workflows, and cross-contamination risks. We evaluated a modified One-Step Multiplex PCR (OS-MPCR) assay designed to streamline library construction while maintaining high analytical sensitivity and diagnostic accuracy. Methods: The OS-MPCR method integrates adapter ligation and library amplification into a single, closed-tube reaction, eliminating intermediate open-tube manipulations. The analytical performance was evaluated using standard reference materials targeting SNVs, Indels, and fusions. Clinical validation was performed on retrospective formalin-fixed paraffin-embedded (FFPE) specimens (DNA, N = 76; RNA, N = 34) from lung cancer patients, using well-established liquid-phase hybrid capture assays as the clinical verification standard. Results: OS-MPCR streamlined the workflow, reducing hands-on time to just 20 minutes. The assay enabled reliable variant detection at 1% frequency with as little as 1 ng of DNA for SNVs/Indels and 5 ng of total RNA for fusions, with low-input quantification limits inherently bounded by absolute molecular copy numbers. In the clinical cohort, OS-MPCR demonstrated 100% diagnostic concordance with the verification method. Quantitative analysis revealed a strong correlation in variant allele frequencies (Spearman r = 0.9777, 95% CI: 0.9668-0.9851, P < 0.0001), with libraries consistently achieving highly reproducible yields and stable, near 1.0 target rates across the entire cohort. Conclusion: OS-MPCR offers a streamlined and low-input alternative to conventional multi-step multiplex PCR methods. By combining reduced hands-on time and intrinsic contamination control with high analytical accuracy, this approach represents a practical refinement for targeted amplicon sequencing in diagnostic workflows. (Copyright: © 2026 Feng et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.) |
| Competing Interests: | I have read the journal’s policy and the authors of this manuscript have the following competing interests: At the time of this research, BL, DS, SW, and YZ were employed by Shanghai Yijian Health Technology Co., Ltd. Shanghai Yijian Health Technology Co., Ltd also provided commercial funding for this research. A patent application related to the library construction methodology reported in this manuscript has been published (Patent title: “A highly efficient and rapid method for constructing specific libraries”; Publication No. CN119979668A), with authors BL and DS listed as inventors. There are no consultancies, products in development, or marketed products to declare. This does not alter our adherence to PLOS ONE policies on sharing data and materials. All other authors declare no competing interests. |
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| Entry Date(s): | Date Created: 20260915 Date Completed: 20260915 Latest Revision: 20260917 |
| Update Code: | 20260917 |
| PubMed Central ID: | PMC13577403 |
| DOI: | 10.1371/journal.pone.0357776 |
| PMID: | 42743120 |
| Βάση Δεδομένων: | MEDLINE |
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