TY - JOUR
T1 - Calibration-free NGS quantitation of mutations below 0.01% VAF
AU - Dai, Peng
AU - Wu, Lucia Ruojia
AU - Chen, Sherry Xi
AU - Wang, Michael Xiangjiang
AU - Cheng, Lauren Yuxuan
AU - Zhang, Jinny Xuemeng
AU - Hao, Pengying
AU - Yao, Weijie
AU - Zarka, Jabra
AU - Issa, Ghayas C.
AU - Kwong, Lawrence
AU - Zhang, David Yu
N1 - Publisher Copyright:
© 2021, The Author(s).
PY - 2021/12/1
Y1 - 2021/12/1
N2 - Quantitation of rare somatic mutations is essential for basic research and translational clinical applications including minimal residual disease (MRD) detection. Though unique molecular identifier (UMI) has suppressed errors for rare mutation detection, the sequencing depth requirement is high. Here, we present Quantitative Blocker Displacement Amplification (QBDA) which integrates sequence-selective variant enrichment into UMI quantitation for accurate quantitation of mutations below 0.01% VAF at only 23,000X depth. Using a panel of 20 genes recurrently altered in acute myeloid leukemia, we demonstrate quantitation of various mutations including single base substitutions and indels down to 0.001% VAF at a single locus with less than 4 million sequencing reads, allowing sensitive MRD detection in patients during complete remission. In a pan-cancer panel and a melanoma hotspot panel, we detect mutations down to 0.1% VAF using only 1 million reads. QBDA provides a convenient and versatile method for sensitive mutation quantitation using low-depth sequencing.
AB - Quantitation of rare somatic mutations is essential for basic research and translational clinical applications including minimal residual disease (MRD) detection. Though unique molecular identifier (UMI) has suppressed errors for rare mutation detection, the sequencing depth requirement is high. Here, we present Quantitative Blocker Displacement Amplification (QBDA) which integrates sequence-selective variant enrichment into UMI quantitation for accurate quantitation of mutations below 0.01% VAF at only 23,000X depth. Using a panel of 20 genes recurrently altered in acute myeloid leukemia, we demonstrate quantitation of various mutations including single base substitutions and indels down to 0.001% VAF at a single locus with less than 4 million sequencing reads, allowing sensitive MRD detection in patients during complete remission. In a pan-cancer panel and a melanoma hotspot panel, we detect mutations down to 0.1% VAF using only 1 million reads. QBDA provides a convenient and versatile method for sensitive mutation quantitation using low-depth sequencing.
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U2 - 10.1038/s41467-021-26308-6
DO - 10.1038/s41467-021-26308-6
M3 - Article
C2 - 34675197
AN - SCOPUS:85117686933
SN - 2041-1723
VL - 12
JO - Nature communications
JF - Nature communications
IS - 1
M1 - 6123
ER -