Firefly: a novel accurate NGS method optimized for cfDNA analysis


Though next-generation sequencing (NGS) technology has greatly advanced our understanding of complex genomic landscapes, limited access to sample material and low signal-to-noise ratios caused by random sequencing errors present significant barriers to realizing its utility.  To address these challenges we have developed Firefly, a novel NGS method uniquely capable of detecting low-frequency variants with high precision in plasma cfDNA.  With Firefly, denatured double-stranded cfDNA is circularized and converted into long tandem repeats using rolling-circle amplification which enable consensus-based concatemer error correction.  Our novel protocol offers several advantages.  (1) Single-stranded ligation results in a high conversion rate.  (2) Repetitive amplification of the original template minimizes the proliferation of errors that can be commonly acquired by regular PCR.  (3) Tandem repeats in concatemers make the detection of authentic mutations accurate and economical.  Performance validation was done using various sample types including: multi-person SNP mixtures, cfDNA simulate, and patient samples collected from individuals with and without cancer.  Firefly revealed 0.1% of variants in 20ng samples of cfDNA at a detection rate of 90% and exhibited an overall error rate of 1 in 1 million.  In summary, we have developed Firefly, a novel method for detecting low-frequency variants with high precision from low input, to facilitate the utilization of plasma cfDNA by both scientists and clinicians.

> Firefly™技术:如暗夜中一点萤火,照亮人类攻克癌症的梦想 <