# Novel Repair Replicase

> **NIH NIH R43** · VARIGEN BIOSCIENCES CORPORATION · 2021 · $399,846

## Abstract

PROJECT SUMMARY
Cytosine deamination of DNA prior to and during amplification, library preparation, and sequencing is the largest
source of errors (C to A) in next-generation sequencing (NGS) data. Thermocycling conditions during PCR sig-
nificantly accelerates cytosine deamination. These errors impede the detection of low-abundance variants such
as driver mutations for small primary and secondary tumors and lower fidelity for complex samples such as
microbiomes and are particularly acute for damaged samples such as formalin-fixed, paraffin-embedded (FFPE)
tissue samples. The NGS-based molecular diagnostics market alone is estimated to reach $2.3B by 2025 and
needs better enzymes.
We have discovered a new group of thermostable proofreading DNA polymerase A enzymes with uracil-DNA
glycosylase (UDG) activity (UDG-DNAP). Preliminary data shows that one of these enzymes, UP19, possesses
very strong 3’ exonuclease activity, UDG activity for removing uracil from DNA, and is capable of robust PCR
amplification under normal thermocycling conditions. In this proposal, we will further develop this enzyme with a
goal of commercializing a first-in-class, thermostable proofreading DNA polymerase A with intrinsic UDG activity
for correcting uracil mistakes in DNA. Specific Aim 1 seeks to characterize and optimize UP19 in standard PCR
reactions against industry standard control enzymes. Specific Aim 2 seeks to measure the fidelity of UP19
against industry standard enzymes using an Illumina platform and high-quality DNA from flash-frozen tumor
samples as the template. Specific Aim 3 seeks to measure uracil error correction of UP19 using FFPE treated
samples and the resulting sequence error rate against industry standard enzymes and a standalone mesophilic
UDG enzyme. As a result of this project, we will have a better understanding of this novel enzyme family and
better understand the applicability of UP19 for NGS applications. The ultimate goal will be to develop a better
enzyme for the NGS market that will reduce error rates and misdiagnoses.
1

## Key facts

- **NIH application ID:** 10323823
- **Project number:** 1R43HG012181-01
- **Recipient organization:** VARIGEN BIOSCIENCES CORPORATION
- **Principal Investigator:** Brian P Hedlund
- **Activity code:** R43 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $399,846
- **Award type:** 1
- **Project period:** 2021-08-10 → 2023-07-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10323823

## Citation

> US National Institutes of Health, RePORTER application 10323823, Novel Repair Replicase (1R43HG012181-01). Retrieved via AI Analytics 2026-05-26 from https://api.ai-analytics.org/grant/nih/10323823. Licensed CC0.

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