The Application of RPA-PfAgo Technology Combined with Multidimensional Data Analysis in the Rapid Detection of the MTHFR A1298C Polymorphism

Abstract

This study presents a novel approach that integrates recombinase polymerase amplification (RPA) with PfAgo protein technology for the rapid and precise detection of the MTHFR A1298C polymorphism. Although traditional genotyping methods are effective, they are often limited by complexity, high cost, and the need for specialized equipment. The RPA-PfAgo technique harnesses the swift isothermal amplification of RPA and the high specificity and sensitivity of PfAgo-mediated DNA cleavage, completing the entire process from sample collection to detection within 90 minutes. The utility of this method has been substantiated through a battery of optimization experiments, parameter analysis, and assessments of sensitivity, specificity, and repeatability, along with clinical validation using oral mucosal samples. The findings indicate that this new technology not only substantially reduces detection time and cost but also offers an effective tool for personalized medicine and disease prevention with high accuracy and reliability.