Rapid recombinase polymerase amplification-based detection of the bar gene for environmental monitoring of genetically modified soybean and rice
TL;DR
A real-time recombinase polymerase amplification (RPA) assay targeting the bar gene for the rapid detection of GM soybean and rice, combined with a simplified NaOH-based DNA extraction method for field applications.
Abstract
ABSTRACT Rapid and reliable methods are required for environmental monitoring of unintended releases of genetically modified (GM) crops. We developed a real-time recombinase polymerase amplification (RPA) assay targeting the bar gene for the rapid detection of GM soybean and rice, combined with a simplified NaOH-based DNA extraction method for field applications. Primer–probe set 4 provided the most specific and reliable detection, while soybean lectin and rice actin served as internal positive controls. Lectin and actin were consistently detected in GM and non-GM samples, respectively. A 0.1 N NaOH extraction method produced DNA of sufficient quality for direct real-time RPA without additional purification. The integrated assay specifically detected the bar gene in GM samples, with no amplification observed in non-GM samples, and generated results within approximately 25 min at 39 °C. This assay provides a rapid, simple, and field-deployable approach for the detection and environmental monitoring of GM soybean and rice.