Loop-mediated isothermal amplification of economically important Ceratitis species (Diptera: Tephritidae)

Abstract

Ceratitis is an economically important genus of fruit flies that originated in Africa, has a wide host range, and causes serious economic losses due to its invasive damage. As a result, it is critical to identify them accurately and quickly in the world. Loop-mediated isothermal amplification (LAMP), as one of the representatives of isothermal amplification technology, has been widely used in the rapid nucleic acid detection of human pathogens and has shown its advantages in the identification of insect agricultural pests. In this study, using the mitochondrial cox1 and cob genes as target genes, the rapid molecular identification of the Ceratitis FARQ complex, C. cosyra, and C. capitata was realized based on LAMP. The experimental conditions optimization results showed that F3/B3:FIP/BIP = 1:8 was the optimal primer concentration ratio and 63 °C was the optimal reaction temperature. The sensitivity of the primers obtained in this study can reach up to 0.01 ng/μl DNA. A loop-mediated isothermal amplification identification technology system was established based on rapid, rough DNA extraction and visual detection of Ceratitis economically important fruit flies. The positive reaction system changed from pink to khaki by visual detection. The identification flow can be completed within 1 hour, including sample processing, DNA extraction, and LAMP visual detection.

Keywords: Ceratitis; identification; loop-mediated isothermal amplification.