Method for detecting Xanthomonas oryzae pv.oryzae, Xoo, X.oryzae pv. Oryzicola, Xoox, X.axonopodis pv. Citri, Xac, and X.campestris pv.campestris, Xcc

A technology for cabbage black rot and rice bacterial blight, applied in the field of molecular biology, can solve problems such as easy pollution, low sensitivity, and cumbersome operation steps

Inactive Publication Date: 2010-12-22
SHENZHEN AUDAQUE DATA TECH +1
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  • Description
  • Claims
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Problems solved by technology

Physiological and biochemical methods, the operation steps are cumbersome, and it takes about 15 days to identify one kind of bacteria, which is too time-consuming, and the relevant personnel need rich experience
Based on the molecular biology of PCR technology, there...

Method used

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  • Method for detecting Xanthomonas oryzae pv.oryzae, Xoo, X.oryzae pv. Oryzicola, Xoox, X.axonopodis pv. Citri, Xac, and X.campestris pv.campestris, Xcc
  • Method for detecting Xanthomonas oryzae pv.oryzae, Xoo, X.oryzae pv. Oryzicola, Xoox, X.axonopodis pv. Citri, Xac, and X.campestris pv.campestris, Xcc
  • Method for detecting Xanthomonas oryzae pv.oryzae, Xoo, X.oryzae pv. Oryzicola, Xoox, X.axonopodis pv. Citri, Xac, and X.campestris pv.campestris, Xcc

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Embodiment 1

[0074] a. DNA extraction: culture test sample A on nutrient agar medium at 30°C for 2-3 days; extract genomic DNA of the test sample to prepare a DNA solution;

[0075] The extraction of bacterial genomic DNA is a known technology, and the specific operation is as follows in the present embodiment: 1) 1.5 mL of bacterial liquid is centrifuged at room temperature at 13000 r / min for 5 min, and the supernatant is removed; Suspended bacteria liquid, 37°C, stand for 2h; 3) Add 385μL of TE, 15μL of 20% SDS, boil for 10min; 4) Extract with an equal volume of phenol chloroform, shake and mix well, centrifuge at 13000r / min at 4°C for 5min, and Transfer the supernatant to a sterilized centrifuge tube; 5) Add 1 mL of ice-free ethanol, and place at -20°C for more than 30 minutes to precipitate DNA; 6) Centrifuge at 13,000 r / min at 4°C for 10 minutes, discard the supernatant, and wash once with ice-absolute ethanol. Wash twice with 75% ethanol, dissolve in 100 μL ddH2O, and store at -20°C ...

Embodiment 2

[0105] Example 3

Embodiment 3

[0107] Example 4

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Abstract

The invention discloses a method for detecting Xanthomonas oryzae pv.oryzae, Xoo, X.oryzae pv. Oryzicola, Xoox, X.axonopodis pv. Citri, Xac, and X.campestris pv.campestris, Xcc. The method comprises the following steps: carrying out PCR (polymerase chain reaction) amplification on the DNA template of a sample to be detected by using two pairs of primers at the same time; then, hybridizing the PCR products with the specific probe arranged on a gene chip; and judging whether the sample to be detected is with Xanthomonas oryzae pv.oryzae, Xoo, X.oryzae pv. Oryzicola, Xoox, X.axonopodis pv. Citri, Xac, and X.campestris pv.campestris, Xcc according to the fact whether the hybridization result is positive or not. Therefore, the method of the invention is capable of detecting four bacteria by one time with high detection speed and has the advantages of direct-viewing and clear detection result and easy judgment.

Description

technical field [0001] The invention belongs to the field of molecular biology, and in particular relates to rapid detection techniques for rice bacterial blight, rice bacterial streak, citrus canker and cabbage black rot. Background technique [0002] The genus Xanthomonas has a wide variety of pathogenicity. The second edition of "Bergey's Handbook of Systematic Bacteriology" included 20 species, 70 pathogenic species with confirmed taxonomic status and 70 taxonomic status not yet identified pathogenic variants. There are 14 quarantine Xanthomonas species included in the current "List of Imported Plant Quarantine Pests of the People's Republic of China". Xanthomonas oryzae pv. oryzae (Xoo) and X. oryzae pv. oryzicola (Xooc) are important bacterial diseases of rice. Rice bacterial blight can cause rice yield reduction by 10% to 30%, and in severe cases it can reach more than 50%. Bacterial spot of rice can cause rice yield reduction by 5% to 10%, and even up to 20% in se...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12Q1/04C12N15/11
Inventor 龙海李一农李芳荣郑耘
Owner SHENZHEN AUDAQUE DATA TECH
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