Multiple PCR primers for detecting lily viruses and application of multiple PCR primers in RNA extraction-free rapid detection method of lily viruses

Abstract

The invention discloses multiple PCR primers for detecting lily viruses. The lily viruses are cucumber mosaic virus (CMV), lily symptomless virus (LSV) and lily mottle virus (LMoV) respectively. Meanwhile, the invention further provides application of the multiple PCR primers in RNA extraction-free rapid detection method of lily viruses. The multiple PCR primers have the advantages that firstly, an RNA extraction method is simplified, a dissecting needle is adopted for picking trace subcutaneous tissues of lily and rapidly transferring the trace subcutaneous tissues to a reverse transcriptionreaction solution, a sampling method is simple, convenient and easy to operate, intermediate links capable of causing RNA degradation and pollution are omitted, an RNA extraction kit is not needed, the detection cost is reduced, and the working efficiency is improved; and secondly, lily virus CMV, LSV and LMoV primers are independently designed and developed, three pairs of specific primers with clear PCR product electrophoresis bands and good repeatability are screened out, amplified fragments are recycled for sequencing, the sequence alignment homology reaches 98% or above, it is proved thatgene fragments obtained through amplification of the primers designed and developed through the method are lily CMV, LSV and LMoV indeed, and it is guaranteed that a detection result is true and reliable.

Description

technical field [0001] The invention relates to the technical field of virus detection, in particular to multiple PCR primers for detecting lily virus and its application in a rapid detection method for lily virus without RNA extraction. Background technique [0002] Lanzhou lily is the only sweet lily in my country's lily varieties, and it is one of the most distinctive famous and special agricultural products in Lanzhou. In recent years, the planting scale of Lanzhou lily has continued to expand, but the production of lily bulbs is propagated through seed bulbs, scale cuttings, etc. Long-term asexual reproduction can easily cause the spread and accumulation of viral diseases, resulting in lower yields, poor quality and lower prices for lilies. Large fluctuations, etc., seriously restricted the industrialization of Lanzhou lily. According to the survey, the main viruses that harm Lanzhou lily are cucumber mosaic virus (CMV), lily asymptomatic virus (LSV), and lily mottle v...

AI Technical Summary

Problems solved by technology

Usually, there are "three highs" in plant tissue RNA isolation, that is, high cost of extraction reagents, high technical difficulty in operation, high requirements for experimental conditions, etc., high cost of RNA extraction kits and cumbersome method steps, time-consuming and laborious, and 30 samples are tested for 3 types The virus takes 2-3 working days; secondly, the extraction of RNA has extremely strict requirements on laboratory conditions and operator skills, and ubiquitous RNase will degrade RNA at any time, resulting in RNA extraction failure or poor quality, which cannot meet the experimental requirements In addition, the specific primer sequence of lily virus is very important for PCR amplification. There are some reports on the detection of lily virus in Lanzhou, but the direct application of previous research results cannot achieve the expected effect

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