By Diagnostics World Staff
August 19, 2025 | Ultima Genomics published updates on the single-nucleotide variant (SNV) detection capabilities of its ppmSeq technology and demonstrated ctDNA detection limits for ppmSeq, which extend beyond the limits of currently available minimal residual disease assays. The updates were published last week at bioRxiv (DOI: 10.1101/2025.08.11.669689).
The work was led by Dan A. Landau, MD, PhD, a core member at the New York Genome Center, and Professor of Medicine & Professor of Physiology and Biophysics at Weill Cornell Medicine lab at the New York Genome Center; Landau is the senior author on the paper.
“State-of-the-art molecular de-noising approaches for DNA sequencing rely on duplex sequencing, where both strands of a single DNA molecule are sequenced to discern true variants from errors arising from single stranded DNA damage,” the authors explain in the paper. “However, such duplex approaches typically require massive over-sequencing to overcome low capture rates of duplex molecules. To address these challenges, we introduce paired plus-minus sequencing (ppmSeq) technology, in which both DNA strands are partitioned and clonally amplified on sequencing beads through emulsion PCR. In this reaction, both strands of a double-stranded DNA molecule contribute to a single sequencing read, allowing for a duplex yield that scales linearly with sequencing coverage across a wide range of inputs.”
Earlier this year the Landau lab published a Nature Methods paper demonstrating the advantages of identifying SNVs with Ultima's flow-based sequencing by synthesis technology versus conventional SBS chemistry (DOI: 10.1038/s41592-025-02648-9). In this new preprint, Landau’s group used ppmSeq, a technology developed by Ultima and native to the Ultima platform, to further improve upon the SNV detection limits. Building on Ultima’s ultra-low error, flow-based sequencing, ppmSeq encodes both strands of DNA molecules in a single sequencing read to enable up to part-per-ten million (10-7) accuracy (SNVQ70) for SNV calling. This exceptional accuracy provides extreme assay sensitivity for low frequency alleles while also requiring 10- to 100-fold less sequencing depth than other error correction techniques.
"With its unique combination of low-cost and high accuracy, ppmSeq marks a real breakthrough in sequencing technology,” said Landau in a press release. “It opens new horizons in studying the somatic genome as a novel frontier in human genetics, and holds enormous promise for clinical applications, including sensitive detection of cancer residual disease."
Highlights from the paper include:
- ppmSeq demonstrated ultrasensitive SNV detection with error rates down to 0.8 x 10-7 for genomic DNA and cell-free DNA.
- ppmSeq demonstrated superior double-stranded DNA (dsDNA) recovery rates compared to standard duplex protocols, reducing sequencing requirements by 10- to 100-fold and enabling a cost-effective whole genome approach.
- The ppmSeq workflow was shown to be highly efficient, yielding greater than 20x coverage per ng of cfDNA, reducing the amount of input material required.
- ppmSeq enabled tumor-informed circulating tumor DNA (ctDNA) detection of 10-5 across multiple cancers, and up to 10-7 in cancers with high mutation burden at 30x sequencing depth, which extends below the limits of current MRD assays.
- ppmSeq identified disease-specific signal in plasma cell-free DNA without the need for a matched tumor.
The authors did note limitations to ppmSeq. Because ppmSeq is a PCR-free library preparation method that relies on hydrogen bonds to carry both DNA strands into a single emulsion, it is limited to whole genome assay. “In addition,” the authors note, “ppmSeq error rates are slightly higher than duplex sequencing methods, likely because traditional duplex sequencing relies on the consensus between two or more independently sequenced molecules to call variants, whereas dsDNA strands are sequenced from a single cluster in ppmSeq. The tradeoff between error rate and yield is therefore an important consideration and is likely to be application dependent.”
But on the whole, the researchers conclude that ppmSeq is a highly accurate and cost-effective option for emerging clinical applications including tumor-informed MRD, tumor-naïve MRD and an opportunity to explore new whole-genome applications in cancer genomics, including somatic mosaicism.