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Duplex PCR (polymerase chain reaction) method for synchronously detecting Burkholderia gladioli pv.alliicola and Erwinia chrysanthemi as quarantine bacteria

A technology for simultaneous detection of rot bacteria, applied in the field of bioengineering, to achieve the effects of strong stability, improved port detection rate and customs clearance efficiency, and improved quarantine level and work efficiency

Inactive Publication Date: 2018-01-26
LIANYUN PORT IMMIGRATION INSPECTION & QUARANTINE BUREAU PEOPLES REPUBLIC OF CHINA
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Problems solved by technology

However, the above studies only detected one type of bacteria, and the molecular detection method for the simultaneous detection of the above two types of bacteria has not been reported yet.

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  • Duplex PCR (polymerase chain reaction) method for synchronously detecting Burkholderia gladioli pv.alliicola and Erwinia chrysanthemi as quarantine bacteria
  • Duplex PCR (polymerase chain reaction) method for synchronously detecting Burkholderia gladioli pv.alliicola and Erwinia chrysanthemi as quarantine bacteria
  • Duplex PCR (polymerase chain reaction) method for synchronously detecting Burkholderia gladioli pv.alliicola and Erwinia chrysanthemi as quarantine bacteria

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Embodiment Construction

[0031] In this study, specific primers were designed based on the ITS sequence of the ribosomal gene of the onion rot fungus, and combined with the published specific primers of the chrysanthemum rot fungus to form a double PCR system to simultaneously detect the onion rot fungus and chrysanthemum rot fungus, in order to improve the The detection rate of quarantine bacterial outbreaks at designated ports for seedlings.

[0032] 1 Materials and methods

[0033] 1.1 Test strains

[0034] The tested onion rot bacteria, chrysanthemum rot bacteria and their related bacterial strains are shown in Table 1.

[0035] 1.2 Reagents, consumables and instruments

[0036] 2x Taq Master Mix and bacterial genome extraction kit were purchased from Shanghai Jierui Biological Company, and DNAMarker was purchased from Dalian Bao Biological Company. The instruments used in the research include PCR instrument (ABI), spectrophotometer (Biochrom), high-speed centrifuge (Eppendorf), gel imaging sys...

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Abstract

The invention discloses a duplex PCR (polymerase chain reaction) method for synchronously detecting Burkholderia gladioli pv.alliicola and Erwinia chrysanthemi as quarantine bacteria. In order to establish the duplex PCR method for synchronously detecting Burkholderia gladioli pv.alliicola and Erwinia chrysanthemi as the quarantine bacteria, a pair of specific primers B3 / B6 is designed, the designed primers are bound with specific primers ADE1 / ADE2 for detecting Erwinia chrysanthemi, a duplex PCR system is built through condition optimization, and specificity and sensitivity verification is performed. With the application of the duplex PCR system, specific bands can be obtained from genomic DNA of Burkholderia gladioli pv.alliicola and Erwinia chrysanthemi and artificial bacteria-bearing samples by amplification. Results indicate that the duplex PCR method can detect Burkholderia gladioli pv.alliicola and Erwinia chrysanthemi simultaneously and can be applied to detection of two typesof quarantine bacteria of tulip bulbs imported from ports.

Description

technical field [0001] The invention belongs to the technical field of bioengineering, and in particular relates to a double PCR method for synchronously detecting the quarantine bacteria onion rot fungus and chrysanthemum rot fungus. Background technique: [0002] Tulips are commonly used in flower exhibitions, potted ornamental and fresh cut flower cultivation. Because of its neat plants, rich colors, beautiful and elegant, it is deeply loved by people. Tulips are highly susceptible to soft rot during the production process. Burkholderia gladioli pv. alliicola and Erwinia chrysanthemi are important bacteria causing tulip soft rot. These two pathogens have been listed as entry phytosanitary pests by China's entry-exit agency because of their wide distribution abroad, high risk of colonization, and great economic harm. In recent years, with the rapid development of the economy and the gradual improvement of people's living standards, domestic flower consumption is showing...

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Application Information

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IPC IPC(8): C12Q1/689C12Q1/686C12Q1/04C12N15/11
Inventor 杨万风邵沛泽谌运清
Owner LIANYUN PORT IMMIGRATION INSPECTION & QUARANTINE BUREAU PEOPLES REPUBLIC OF CHINA
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