{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,15]],"date-time":"2026-09-15T18:04:23Z","timestamp":1789495463737,"version":"build-2803163510"},"reference-count":32,"publisher":"Public Library of Science (PLoS)","issue":"9","license":[{"start":{"date-parts":[[2026,9,15]],"date-time":"2026-09-15T00:00:00Z","timestamp":1789430400000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"Medical and Health Technology Development Program in Shandong Province","award":["202301040579"],"award-info":[{"award-number":["202301040579"]}]}],"content-domain":{"domain":["www.plosone.org"],"crossmark-restriction":false},"short-container-title":["PLoS One"],"abstract":"<jats:sec id=\"sec001\">\n                    <jats:title>Background<\/jats:title>\n                    <jats:p>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.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec id=\"sec002\">\n                    <jats:title>Methods<\/jats:title>\n                    <jats:p>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\u2009=\u200976; RNA, N\u2009=\u200934) from lung cancer patients, using well-established liquid-phase hybrid capture assays as the clinical verification standard.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec id=\"sec003\">\n                    <jats:title>Results<\/jats:title>\n                    <jats:p>\n                      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\n                      <jats:italic>r<\/jats:italic>\n                      \u2009=\u20090.9777, 95% CI: 0.9668\u20130.9851,\n                      <jats:italic>P<\/jats:italic>\n                      \u2009&lt;\u20090.0001), with libraries consistently achieving highly reproducible yields and stable, near 1.0 target rates across the entire cohort.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec id=\"sec004\">\n                    <jats:title>Conclusion<\/jats:title>\n                    <jats:p>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.<\/jats:p>\n                  <\/jats:sec>","DOI":"10.1371\/journal.pone.0357776","type":"journal-article","created":{"date-parts":[[2026,9,15]],"date-time":"2026-09-15T17:27:08Z","timestamp":1789493228000},"page":"e0357776","update-policy":"https:\/\/doi.org\/10.1371\/journal.pone.corrections_policy","source":"Crossref","is-referenced-by-count":0,"title":["A novel one-step multiplex PCR method (OS-MPCR) for efficient NGS library preparation in cancer"],"prefix":"10.1371","volume":"21","author":[{"given":"Haiyan","family":"Feng","sequence":"first","affiliation":[{"name":"Zibo First Hospital","place":["Zibo, Shandong, China"],"department":["Department of 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