Molecular Diagnosis Enabled by Dynamic DNA Assembly and CRISPR Cas Toolbox

The accurate and timely detection of disease biomarkers at the point-of-care is essential to ensure effective treatment and prevention of diseases. CRISPR-based diagnostics (CRISPR-Dx) is revolutionizing molecular diagnosis by offering exceptional sensitivity, specificity, and simplicity. Research in our laboratory focuses on leveraging CRISPR-Dx and dynamic DNA assembly to create better analytical tools for diagnosing both human and crop diseases. In this talk, I will first introduce a plasmonic CRISPR Cas12a assay that combines the trans-cleavage activity of Cas12a and DNA functionalized gold nanoparticles for the visual, colorimetric detection of grapevine viral infections. I will then describe a molecular translator approach that expands CRISPR-Dx to protein-based biomarkers. Such CRISPR-based ultrasensitive protein assays enabled the detection of attomolar concentrations of SARS-CoV-2 antibodies in serum samples collected from vaccinated kidney transplant recipients who are shown to produce “undetectable” anti-SARS-CoV-2 using standard immunoassay. Finally, I will describe the development of a CRISPR-like assay using RNA-cleaving DNAzyme coupled with isothermal sequence and signal amplification, which demonstrated enhanced accessibility and multiplexity over CRISPR-DX.

Learning Objectives:

1. Discuss possible strategies to enable colorimetric readout for CRISPR diagnostics.

2. Evaluate design approaches that enable CRISPR diagnostics for protein-based biomarkers.

3. Discuss possible roles of CRISPR diagnostics in the face of emerging infectious diseases such as COVID-19.