Establishment of Microfluidic Chip Method for Rapid Screening of <i>Tobacco Rattle Virus</i>

Abstract

<i>Tobacco rattle virus</i> (TRV), member of the genus <i>Tobravirus,</i> is an important plant pathogen with a wide host range beyond any known plant viruses. It is able to be transmitted by nematodes of the genera <i>Trichodorus</i> and <i>Paratrichodorus</i> (Trichodoridae). In this paper, a rapid screening method was established for inspection of TRV in seeds or other plant materials, combining microfluidic chip technique and fluorescence detection system. The particular region (located at 496-995) of TRV strain (accession number: KP100069.1) was adopted for primer design, method establishment and improvement. Through analysis, this method was proved to be specific for only detecting sample infected by TRV, while 7 other leaf materials infected by <i>Tomato spot wilt virus</i> (TSWV), <i>Tobacco ringspot virus</i> (TRSV), <i>Cucumber mosaic virus</i> (CMV), <i>Lily symptomless virus</i> (LSV), <i>Lily mosaic virus</i> (LMV), <i>Tomato ringspot virus</i> (ToRSV), and <i>Prunus necrotic ringspot virus</i> (PNRSV), respectively, showed negative results. Sensitivity tests represented a detection limit as low as 1.00×10²copies/μL which is beyond normal PCR assays. In one word, microfluidic chip based analyzing platforms hold high promises to enable high-throughput and high-precision screening with less sample consumption, fast detection, simple operation, multi-functional integration, small size, multiplex detection and portability, thus promoting the development of biosecurity diagnostics.