关键词: 柑橘脉突病毒, 实时荧光定量RT-PCR

Abstract:

In order to fast by detect Citrus vein enation virus（CVEV）with sensitivity and specificity，a Reverse Transcription Quantitative Polymerase Chain Reaction（RT-qPCR）assay was developed with selective primer pairs（EVqF4/EVqR4）and optimized conditions. Using the method，only sample infected with CVEV showed positive，while those with other five citrus pathogens were negative. The sensitivity of the method was 100 times higher than the conventional RT-PCR. There was a good linear（R² = 0.992）relationship between the threshold cycle and CVEV template concentration，while the amplification efficiency of the RT-qPCR was 101.8%. The coefficients of variation of the intra- and inter-assay were both within 2.85%，indicating a good reproducibility of the method. The results also showed that CVEV was uneven distributed in sour orange plants and hybrid citrus plants，and the amount of the virus in roots was the highest（162.52 and 45.32 copies · ng^-1 RNA，respectively），followed by that in barks and leaves.

Key words: Citrus vein enation virus（CVEV）, quantitative RT-PCR, application