Interlaboratory Validation Study of an Event-Specific Real-time Polymerase Chain Reaction Detection Method for Genetically Modified 55-1 Papaya

Abstract

Genetically modified (GM) papaya line 55-1 (55-1) is resistant to papaya ringspot virus infection, and is commercially available in several countries. A specific detection method for 55-1 is required for mandatory labeling regulations. An event-specific real-time PCR method was developed by our laboratory. To validate the method, interlaboratory validation of event-specific qualitative real-time PCR analysis for 55-1 was performed in collaboration with 12 laboratories. DNA extraction and real-time PCR reaction methods were evaluated using 12 blind samples: six non-GM papayas and six GM papayas in each laboratory. Genomic DNA was highly purified from all papayas using an ion-exchange column, and the resulting DNA sample was analyzed using real-time PCR. Papaya endogenous reference gene chymopapain (CHY) and the event-specific 55-1 targeted sequence were detected in GM papayas whereas CHYalone was detected in non-GM papayas in all laboratories. The cycle threshold values of CHYand the 55-1 targeted sequence showed high repeatability (RSD, 0.6-0.8%) and reproducibility (RSDR 2.2-3.6%). This study demonstrates that the 55-1 real-time PCR detection method is a useful and reliable method to monitor 55-1 papaya in foods.