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Use of Reverse Transcription Loop‐Mediated Isothermal Amplification for the Detection of Z ucchini yellow mosaic virus
Author(s) -
Kuan ChengPing,
Deng TingChin,
Huang HungChang,
Chi HsiangHui,
Lu YiLin
Publication year - 2014
Publication title -
journal of phytopathology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.53
H-Index - 60
eISSN - 1439-0434
pISSN - 0931-1785
DOI - 10.1111/jph.12175
Subject(s) - loop mediated isothermal amplification , zucchini yellow mosaic virus , biology , sybr green i , squash , reverse transcription loop mediated isothermal amplification , microbiology and biotechnology , virology , reverse transcription polymerase chain reaction , real time polymerase chain reaction , reverse transcriptase , virus , cucumber mosaic virus , plant virus , polymerase chain reaction , horticulture , dna , gene , biochemistry , messenger rna
A Reverse Transcription Loop‐Mediated Isothermal Amplification ( RT ‐ LAMP ) assay was employed to develop a simple and efficient system for the detection of Z ucchini 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.