# Integrated Instrument for non-natural aptamer generation

> **NIH NIH R01** · STANFORD UNIVERSITY · 2020 · $314,000

## Abstract

Project Summary
DNA and RNA aptamers are a useful class of synthetic affinity reagents. However, their performance can be
greatly improved through the site-specific incorporation of chemically modified, ‘non-natural’ nucleotides that
provide a greater chemical repertoire to enable superior aptamer affinity and specificity. Because a broad
spectrum of chemical functional groups can be incorporated, non-natural aptamers offer the exciting potential
for targeting molecules for which the generation of monoclonal antibodies remains difficult, such as small-
molecule drugs, metabolites and carbohydrates. Unfortunately, the access to non-natural aptamers is severely
limited. This is because the process of generating non-natural aptamers is technically challenging and limited
to a few specialized laboratories. The goal of this project is to develop an integrated instrument, the Non-Natural
Aptamer Array (N2A2) that eliminates these bottlenecks and enable rapid and facile non-natural aptamer
discovery at virtually any research laboratory. The N2A2 will be built on a modified version of a benchtop
commercial sequencer (Illumina MiSeq), and will perform every stage of non-natural aptamer discovery—
including sequencing, screening and binding measurements—as part of a single work-flow. There are three main
innovative aspects of our N2A2 system. First, our approach will entirely eliminate the need for polymerase
engineering, and thus allows us to incorporate virtually any chemical functional group through click chemistry.
Second, N2A2 will enable us to directly obtain the binding affinity (Kd) of ~10^7 aptamers directly in complex
samples (e.g. cell lysate or serum), thereby resulting in aptamers with high-specificity. Finally, we will develop
a machine-learning (ML) approach to identify key motifs (“k-mers”) and predict novel sequences with potentially
higher affinity and specificity that can be tested using the N2A2 instrument. We believe this powerful combination
of massively parallel, sequence-linked binding measurements with ML-based predictions will allow us to explore
sequence space that is currently inaccessible to traditional in vitro selection methods, and enable us to discover
aptamers with superior performance. The success of this project will produce an integrated instrument that
greatly streamlines and accelerates the discovery of non-natural aptamers for a wide range of targets in complex
media. The instrument is based on a commercially available sequencer and we will make all software available
to the public. In this way, we believe the N2A2 instrument could broadly expand access to robust, high quality,
custom affinity reagents for biomedical research and clinical diagnostics.

## Key facts

- **NIH application ID:** 9872178
- **Project number:** 5R01GM129313-03
- **Recipient organization:** STANFORD UNIVERSITY
- **Principal Investigator:** HYONGSOK Tom SOH
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $314,000
- **Award type:** 5
- **Project period:** 2018-06-01 → 2022-02-28

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9872178

## Citation

> US National Institutes of Health, RePORTER application 9872178, Integrated Instrument for non-natural aptamer generation (5R01GM129313-03). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/9872178. Licensed CC0.

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