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Polymerase chain reaction (PCR) method for specifically detecting pectobacterium carotovorum

A technology of carrot soft rot and pectobacillus, which is applied in the directions of microorganism-based methods, biochemical equipment and methods, and microbial determination/inspection, etc., can solve problems such as poor bacterial specificity, and achieve high specificity.

Inactive Publication Date: 2012-07-11
HUNAN AGRICULTURAL UNIV
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Problems solved by technology

[0004] The technical problem to be solved by the present invention is: aiming at the deficiencies of the above-mentioned prior art, a PCR method for specific detection of Pectinobacillus carotovora is provided, which solves the problem of poor specificity in identifying Pectinobacillus bacteria according to the 16S rDNA sequence , can accurately determine whether plant soft rot is caused by Pectin Bacillus carotovora

Method used

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  • Polymerase chain reaction (PCR) method for specifically detecting pectobacterium carotovorum
  • Polymerase chain reaction (PCR) method for specifically detecting pectobacterium carotovorum
  • Polymerase chain reaction (PCR) method for specifically detecting pectobacterium carotovorum

Examples

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Embodiment 1

[0032] Embodiment one detects diseased plant

[0033] Taking artichoke rhizomes as an example, Pectinbacterium carotovora soft rot, which causes artichoke root rot disease, was detected.

[0034] Inoculate the strains isolated from artichoke rhizome rot diseased plants and identified as Pectinobacillus carotovora soft rot on disease-free artichoke stems, and after the initial symptoms appear, extract DNA from the diseased tissue as a PCR template, and then use the present invention to The primer pair was used for PCR, and the PCR parameters were the same as before. PCR results showed that artichoke stems inoculated with P. carotovora were positive, while disease-free artichoke stems not inoculated with P. carotovora were negative, see Figure 5 . It shows that the specific primer can be used to detect the diseases caused by Pectinbacterium carotovora soft rot.

Embodiment 2

[0035] Embodiment two distinguishes similar bacteria

[0036] According to the aforementioned bacterial DNA extraction and PCR methods, 4 kinds of bacteria were tested: Pectinobacillus carotovora, Blackleg of potato, Erchetia chrysanthemi and Pantoea pineapple. Only the sample of Pectinobacillus carotovora was positive, and the other three were positive. Bacterial sample was negative, see Figure 6 .

[0037] Thus, theoretically, the results of BLAST of the L32171.1 sequence and forward and reverse primers, and practically, the results of Examples 1 and 2 all confirm the specificity of the primer pair of the present invention. Using this pair of primers for PCR detection can accurately distinguish Pectinobacillus carotovora from other bacteria, and can distinguish Pectinobacillus carotovora from other bacteria of the genus Pectinbacterium.

[0038]

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Abstract

The invention relates to a polymerase chain reaction (PCR) method for specifically detecting pectobacterium carotovorum. A pair of primers is designed according to a base sequence (GenBank:L32171.1) of a pectobacterium carotovorum subsp. carotovorum No.71 strain pectate lyase gene, and a basic local alignment search tool (BLAST) result proves that the pair of primers has high specificity. When the pair of primers is used for PCR detection, bacteria of other genera can be removed, the pectobacterium carotovorum can be distinguished from other bacteria of pectobacterium, and the specificity is high.

Description

technical field [0001] The invention relates to a method for detecting Pectobacterium carotovorum (Pectobacterium carotovorum, synonymous name Erwinia carotovora), in particular to a PCR (polymerase chain reaction) technique for specifically detecting the bacterium in soft rot of plants. Background technique [0002] Plant soft rot is an important class of plant diseases, which can be caused by fungi or bacteria. The rot of the same plant can be caused by different species of pectin bacteria. For example, potatoes can be affected by three kinds of pectin bacteria, Pectobacterium carotovorum and potato blackleg fungus Pectobacterium atrosepticum (synonym Pectobacterium carotovorum var. atrosepticum, Erwinia carotovora subsp.atroseptica), and Pectobacterium chrysanthemi (synonym Erwinia chrysanthemi), thus causing difficulties in pathogen identification. Some people have introduced the routine identification methods of the above three bacteria in seed potatoes, that is, after...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12Q1/04C12R1/01
Inventor 高必达王晓丽
Owner HUNAN AGRICULTURAL UNIV
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