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Development of Real‐time RT ‐ PCR Assays for Detection and Quantification of Pineapple Mealybug Wilt‐associated Viruses‐1
Author(s) -
Chang KaiWei,
Ho TsingFen,
Lin YuChang,
Kuan ChengPing
Publication year - 2016
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.12462
Subject(s) - mealybug , taqman , real time polymerase chain reaction , biology , reverse transcription polymerase chain reaction , virology , polymerase chain reaction , microbiology and biotechnology , hsp70 , messenger rna , gene , heat shock protein , botany , biochemistry , pest analysis
Pineapple mealybug wilt‐associated virus ‐1 ( PMW aV‐1) is a major causative agent for pineapple disease. A real‐time, one‐step reverse transcription ( RT )–polymerase chain reaction ( PCR ) assay using TaqMan probes was developed for the detection of PMW aV‐1. Real‐time RT ‐ PCR , targeting the amplification of heat‐shock protein 70 homolog ( hsp70 h ) gene fragment, was highly specific and capable of distinguishing PMW aV‐1 from PMW aV‐2 and PMW aV‐3, which are two other common virus species in pineapple plants. Sensitivity assay revealed that real‐time RT ‐ PCR was approximately 100‐ to 1000‐fold more sensitive than standard one‐step RT ‐ PCR , when total RNA of infected plant and PMW aV‐1 hsp70 h transcript were used, respectively. This real‐time RT ‐ PCR assay is rapid, specific and sensitive, which can be used for PMW aV‐1 diagnosis as well as surveillance.