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PCR method for specific detection of watermelon acidovorax citrulli

A kind of fruit spot fungus, specific technology, applied in the field of PCR technology for specific detection of fruit spot fungus on watermelon, can solve the problems such as poor specificity of primers, and achieve high specificity effect

Inactive Publication Date: 2012-06-13
HUNAN AGRICULTURAL UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0007] The technical problem to be solved by the present invention is: aiming at the above-mentioned deficiencies in the prior art, a kind of PCR method for specific detection of fruit spot bacterium (Acidovorax citrulli) on watermelon is provided, which solves the problem of poor specificity of primers in the PCR detection of fruit spot bacterium on watermelon problem, can accurately detect the situation of fruit spot bacteria on watermelon

Method used

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  • PCR method for specific detection of watermelon acidovorax citrulli
  • PCR method for specific detection of watermelon acidovorax citrulli
  • PCR method for specific detection of watermelon acidovorax citrulli

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Embodiment 1

[0040] Five fruit spot strains were obtained from Zhao Tingchang's laboratory of the Institute of Plant Protection, Chinese Academy of Agricultural Sciences (all have done 16S ribosomal RNA gene sequencing, BLAST comparison and inoculation test). First use the primers of the present invention to pair the bacterial strains on 2 watermelons (collected respectively from Hainan Sanya (watermelon bacterial strain 1) and Heilongjiang Harbin (watermelon bacterial strain 2)) and the bacterial strain on 1 muskmelon (collected from Inner Mongolia Linhe City) these 3 A preliminary PCR test was performed for 1 strain and 1 miscellaneous bacteria. As a result, only 2 strains on watermelon were positive, and the amplified band size was consistent with the expected 333 bases. Melons were negative for strains and miscellaneous bacteria, see Figure 5 .

[0041] In order to ensure the accuracy of the test, another 2 melon strains (respectively collected from Xinjiang (melon strain 2) and Hai...

Embodiment 2

[0052] Example 2 Detection of diseased watermelon leaves

[0053] The primer pair of the present invention is used to detect the watermelon leaves infected with fruit spot disease, and the PCR detection procedure is the same as before, but the detection object is not the strain, but the watermelon leaves that are inoculated with the watermelon fruit spot disease bacteria. Results All positive results were obtained from 6 watermelon leaves artificially inoculated with P. Figure 8 ), the size of the amplified band was in line with the expected one.

[0054] Known from above embodiment, this pair of specific primers of the present invention is specific to fruit spot bacterium on watermelon, adopt this primer to do PCR to fruit spot bacterium on watermelon, can accurately distinguish fruit spot bacterium on watermelon and other bacteria accurately Distinguish and can distinguish fruit blotch fungus on watermelon from fruit blotch fungus on melon.

[0055]

[0056]

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Abstract

The invention relates to a PCR (polymerase chain reaction) method for specific detection of watermelon acidovorax citrulli. The method designs a pair of primers according to an assumed IV type pilin gene (GenBank: CP000512.1) sequence on the complete genome sequence of a watermelon acidovorax citrulli strain AAC00-1, and a BLAST result indicates that the pair of primers have very good specificity. During PCR detection, the pair of primers not only can eliminate other bacteria, and can distinguish the acidovorax citrulli in watermelons and the acidovorax citrulli in muskmelons. And the pair ofprimers can distinguish an acidovorax citrulli strain in a watermelon from other bacteria accurately and have high specificity.

Description

technical field [0001] The invention relates to a detection method for Acidovorax citrulli on watermelon, in particular to a PCR (polymerase chain reaction) technique for specifically detecting Acidovorax citrulli on watermelon. Background technique [0002] In terms of PCR detection, the specificity of PCR primers is not satisfactory. Some literatures do not use similar bacteria as controls for PCR or real-time fluorescent PCR, and only introduce sensitivity but not specificity. There are other bacteria such as angular leaf spot bacteria. Some PCR primer designs are very unreasonable. For example, the PCR primer 5'-GACCAGCCACACTGGGAC-3' of a dissertation actually showed that the sequence in the front did not have the sequence of Phytophthora japonicus after BLAST. [0003] The same is true for the PCR primers designed in another master's thesis. Using the designed primers to do BLAST, the results show that there is no P. rotifera sequence in the first 30 sequences. Articl...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12Q1/04
Inventor 高必达
Owner HUNAN AGRICULTURAL UNIV
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