Use of Reverse Transcription Loop‐Mediated Isothermal Amplification for the Detection of Zucchini yellow mosaic virus

Abstract

AbstractA Reverse Transcription Loop‐Mediated Isothermal Amplification (RT‐LAMP) assay was employed to develop a simple and efficient system for the detection of Zucchini yellow mosaic virus (ZYMV) in squash and melon plants. The RT‐LAMP assay took 30 min under isothermal condition at 64°C by employing a set of four primers targeting ZYMV. The sensitivity of RT‐LAMP was 10‐fold greater than that of the RT‐PCR assay in the detection of ZYMV in infected tissues of squash and melon. No reaction was detected from the tissues of healthy plants by either RT‐LAMP or RT‐PCR assay. The RT‐LAMP product of the tested samples can be visualized by staining directly in the tube with SYBR Green I dye. The sensitivity of SYBR Green I staining method is similar to that analyzed by gel electrophoresis. Field‐grown squash and melon plants were tested using RT‐PCR and RT‐LAMP. Both RT‐LAMP and PCR could detect ZYMV in symptomatic or symptomless tissues of infected plants. However, the RT‐LAMP assay is superior to RT‐PCR because it is rapid, simple, and highly sensitive; therefore, RT‐LAMP is a useful and practical method for detection of ZYMV in cucurbits.