Method for synchronously detecting three viruses of lily by immunocapture triple RT-PCR (reverse transcription-polymerase chain reaction)

Abstract

The invention provides a method for synchronously detecting a latent symptom virus, a mottle virus and a cucumber mosaic virus of lily by immunocapture triple RT-PCR (reverse transcription-polymerase chain reaction). The method comprises the following steps: specifically capturing LSV, LMoV and CMV particles by polyclonal antibodies of LSV, LMoV and CMV in the same PCR reaction tube; and carrying out triple RT-PCR and agarose gel electrophoresis detection on a target segment by the specific primers of the LSV, the LMoV and the CMV. According to the method, by combination of ELISA and RT-PCR, existing immunocapture triple RT-PCR is improved; and synchronous detection of three lily viruses LSV, LMoV and CMV by immunocapture triple RT-PCR is achieved, so that the immunocapture RT-PCR technology is perfected.

Description

technical field [0001] The invention relates to a method for synchronous detection of lily recessive virus, mottle virus and cucumber mosaic virus, and further relates to a method for synchronously detecting lily recessive virus, mottle virus and cucumber mosaic virus by using immune capture triple RT-PCR technology method. Background technique [0002] Since lily virus was discovered in the 1940s, it has become a worldwide disease that endangers the production of lily bulbs and the quality of fresh cut flowers; so far, more than 20 kinds of lily viruses have been found to infect lily viruses, including lily hidden virus (Lilysymptomlessvirus, LSV), Lily mottle virus ( LMoV ) and cucumber mosaic virus ( CMV ) are the three most common and serious viruses. [0003] LSV belongs to the genus Carlavirus and is a positive-sense single-stranded RNA virus; the LSV genome is non-segmented RNA, with a total length of 8394bp, a cap structure at the 5′ end, and a polyA tail at the 3′ ...

AI Technical Summary

Problems solved by technology

[0006] The more effective means of preventing and treating lily virus disease is to use non-toxic propagation materials or carry out detoxification and rapid propagation, which depends on sensitive, rapid and reliable detection technology; enzyme-linked immunosorbent assay (ELISA) based on the antibody reaction of virus coat protein antigen And RT-PCR detection based on viral nucleic acid is currently a relatively widely used method for detecting viruses; however, ELISA detects a reaction system that can only complete the detection of a virus, and cannot detect multiple viruses simultaneously; although RT-PCR can detect multiple viruses However, the detection must extract high-quality RNA. The bulbs of lily, narcissus, tulip and other bulbous flowers are rich in polyphenols and polysaccharides. The residual polyphenols and polysaccharides in the extracted RNA will seriously affect the sensitivity and specificity of the detection. Reliability; Immunocapture RT-PCR or IC-RT-PCR combines the advantages of ELISA and RT-PCR with other methods, has higher sensitivity, and is more convenient and efficient. More importantly, it does not need to extract RNA, and the detection cost is low. Affected by polysaccharides and polyphenols, it is very suitable for the virus detection of lily, narcissus, and tulip bulbs; as for immune capture RT-PCR, there have been reports on the detection of various plant viruses by this method, but the established detection systems can only Complete the detection of a single virus, so far there is no report on the simultaneous detection of two or more viruses using this method

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