# High-throughput cell-free production of human genetic variants

> **NIH NIH R43** · SYNVITROBIO, INC. · 2020 · $252,116

## Abstract

Project Summary
The rapidly evolving field of human functional genomics has created the promise of personalized medicine. While
extensive sequencing has generated valuable genetic association data, our ability to test linked variants in
disease has not kept pace. As a result, most Single Nucleotide Polymorphisms (SNPs) are classified as Variants
of Unknown Significance (VUS). Following up on function is essential to determine the causal variants and genes
for disease biology and drug discovery. There are currently no technologies to produce variants en masse from
genomic association data to establish function. We propose using Synvitrobio’s cell-free protein expression and
purification platform to rapidly express and purify human SNP variants for downstream functional testing. Cell-
free systems take only 8 hours to express, rather than days to weeks in cells, since there is no need for cloning
and transformation. They are also at least 10-fold cheaper to run than cells, can be run in high-throughput as
reactions (384 well plates), and can be scaled up for protein purification. In short, our approach provides a
scalable alternative to cell-based heterologous expression. This Phase I study proposes to produce model
human SNP variants using traditional in cellulo methods and compare the specific activity to proteins produced
via Synvitrobio’s cell-free platform. We propose two aims that de-risk the strategy of constructing panels of
human SNP variants using high-throughput cell-free tools. Aim 1 focuses on a class of protein known to express
in E. coli systems, whereas the protein class in Aim 2 is more challenging in typical prokaryotic expression
systems. Building a toolbox for variant expression and purification would facilitate understanding the causal
variants and genes for disease biology and drug discovery.

## Key facts

- **NIH application ID:** 10155618
- **Project number:** 1R43HG011602-01
- **Recipient organization:** SYNVITROBIO, INC.
- **Principal Investigator:** Zachary Z. Sun
- **Activity code:** R43 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $252,116
- **Award type:** 1
- **Project period:** 2020-09-11 → 2022-03-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10155618

## Citation

> US National Institutes of Health, RePORTER application 10155618, High-throughput cell-free production of human genetic variants (1R43HG011602-01). Retrieved via AI Analytics 2026-05-25 from https://api.ai-analytics.org/grant/nih/10155618. Licensed CC0.

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