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Updated: Jun 14, 2026
Rare Event Detection Using Error-corrected DNA and RNA Sequencing
Published on: August 3, 2018
Benchmarking Q40 sequencing for sensitive and efficient detection of rare genomic variants
Shumeng Duan1, Yaqing Liu1, Xiaorou Guo1
1State Key Laboratory of Genetics and Development of Complex Phenotypes, School of Life Sciences and Human Phenome Institute, Fudan University, Shanghai, China.
Background:
Phred quality score (Q score) is critical for sequencing accuracy, yet the impact of Q40-achieving sequencing technologies (99.99% accuracy) on detecting subtle biological variations remains unvalidated.
Results:
Using a comprehensive set of well-established DNA/RNA reference materials (Quartet, NIST-RM8398, SEQC2-HCC1395/BL, MAQC, and ERCC), we benchmarked Q40 sequencing (Element AVITI) against the conventional Q30 standard (Illumina NovaSeq 6000). Q40 reduced required sequencing depth by 33.3% while maintaining accuracy for germline variants (20 × vs. 30 ×) and somatic single-nucleotide variant/insertion-deletion (SNV/InDel) (60 × vs. 90 ×). Crucially, Q40 enhanced sensitivity for low-frequency somatic mutations (variant allele frequency, VAF ≤ 0.2) by 33.3% and sixfold higher copy number variation (CNV) detection reproducibility (60.3% vs. 10.4%) with Q40 at 30 × depth, directly reducing per-sample volumes by 33.3-60% and theoretically reducing sequencing costs by 2.2-31.7%. In addition, Q40 improved the discriminatory resolution between biological samples with 13.1% signal-to-noise ratio (SNR) enhancement.
Conclusions:
Taken together, our findings establish the value of Q40 sequencing as a sensitive and cost-effective method for low-frequency variant detection. While this positions it as a promising tool for precision oncology, its performance in real-world clinical applications remains to be evaluated in future studies.
