Objective A multiplex RT-PCR assay to simultaneously detect citrus yellow vein clearing virus (CYVCV), citrus tristeza virus (CTV), and hop stunt viroid (HSVd) was developed.
Method Multiplex RT-PCR primers were designed, and their specificity was analyzed. The optimal concentration ratio, annealing temperature and sensitivity of primers were determined. Moreover, citrus samples from Fujian region were detected by the established multiplex RT-PCR.
Result The optimized concentration ratio of CYVCV-F/R, CTV-F/R, and HSVd-F/R primers was 1:1:2, and the annealing temperature at 52.9 ℃ for the assay. The assay displayed a sensitivity at 10-2 dilution and the positive detections on CYVCV of 47.1%, on CTV of 56.7%, and on HSVd of 22.9% on 157 detected samples.
Conclusion A rapid, accurate, applicable multiplex RT-PCR method for CYVCV, CTV, and HSVd detections was successfully established.
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