High-throughput analysis of point mutations in maize promoters
A novel platform to assess the impact of single-nucleotide changes in maize promoter regions on gene expression, leveraging a protoplast transient assay and RNA-seq for high-throughput analysis. Initially focused on maize, this system can be adapted to other plant species.
Overview
This innovative research platform decodes the regulatory effects of point mutations in maize promoter regions. It evaluates how single-nucleotide changes affect gene expression, specifically in maize genes involved in phenylpropanoid and flavonoid biosynthesis. Using a protoplast transient assay and high-throughput RNA-seq, this system provides detailed maps of promoter contributions to gene expression and transcription factor-mediated regulation. Although initially applied to maize, the platform's design allows easy adaptation to other plant species, enhancing its versatility and applicability in plant genomics.
Technical specifications
- Methodology: Utilizes a protoplast transient assay combined with high-throughput multiplexed RNA-seq.
- Focus: Evaluates promoter variants of 20 maize genes critical to phenylpropanoid and flavonoid biosynthesis pathways.
- Mutagenesis: Random PCR introduces single-nucleotide changes across 1 kb promoter regions.
- Reporter System: Each mutant is linked to a unique transcribed barcode and cloned upstream of GFP.
- Analysis: mRNA extracted and analyzed ~24 hours post-transformation to determine variant activity.
- Transformation: Libraries co-transformed with specific transcription factors driven by p35S.
Technology readiness level
The technology is at TRL 4, reflecting the completion of validation procedures in laboratory settings. Future work will focus on further refinement and adaptation for broader use across different plant species.
About Michigan State University
Michigan State University is a major public land‑grant research university with a comprehensive academic portfolio and a large research enterprise. Industry partners engage through an on‑campus U.S. Department of Energy national user facility and shared core laboratories with user access. The university provides a chemical process scale‑up pilot plant on Michigan’s lakeshore, a research and technology park, and a Grand Rapids health innovation campus linking researchers with clinical partners. A statewide extension network supports field deployment and workforce training across Michigan’s manufacturing corridor. Research is backed by competitive federal funding from NSF, NIH, DOE, USDA, and DoD, while dedicated tech transfer and corporate engagement teams—supported by an affiliated research foundation—accelerate IP, licensing, startups, and sponsored research.
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