Molecular identification method of trichothecene type-A toxins of fusarium

A fusarium and toxin technology, which is applied in the field of molecular identification of fusarium trichothecenes A toxins, can solve the problems that the toxin-producing species have not been verified by chemical analysis, the results cannot be consistent with each other, and the reliability is not high.

Inactive Publication Date: 2014-06-18
HUAZHONG AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

But so far, there are very few studies on the molecular identification of class A trichothecenes. Nicholson et al. used the Tri4 gene sequence information to detect the production of class A or B toxins in 14 strains of 13 species of Fusarium (Nicholson et al. Detection and differentiation of trichothecene and enniatin-producing Fusarium species on small-grain cereals. Eur. J. Plant Pathol., 2004, 110: 503-514), the results of some strains detected by two pairs of primers cannot be compared with each other coincide, and the types of toxins produced by each strain have not been verified by chemical analysis, so the reliability is not high

Method used

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  • Molecular identification method of trichothecene type-A toxins of fusarium
  • Molecular identification method of trichothecene type-A toxins of fusarium
  • Molecular identification method of trichothecene type-A toxins of fusarium

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0047] Detection and identification of toxin produced by strain Fusarium langsethiae

[0048] After the strain Fusarium langsethiae (see Table 1) was activated and cultured on the PDA plate for 5 days, a 2 cm diameter hyphae block was punched at the edge of the colony with a punch, and then a 3-point inoculation method (Torp and Langseth, Production of T-2 toxinby a Fusarium resembling Fusariumpoae. Mycopathologia, 1999, 147: 89-96) evenly inoculate the mycelial mass into T-2 toxin induction medium (recipe: yeast extract 20g, sucrose 150g, MgSO 4 ·7H 2 O 0.5g, 20g agar, add distilled water to make the volume to 1L, sterilize at 120°C for 20min before use) on the plate. After the parafilm was sealed, cultured in the dark at 25°C for 14 days. After the culture was ground and crushed by a stirrer, 5 g of the sample was weighed into a 50 ml Erlenmeyer flask, and then 25 ml of acetonitrile-water (volume ratio 84:16) was added, and the culture was shaken and extracted at 150 r / min for ...

Embodiment 2

[0052] Detection and identification of toxin produced by Fusarium poae

[0053] After the strain Fusariumpoae (see Table 1) was activated and cultured on the PDA plate for 5 days, the hyphae block with a diameter of 2 cm was punched at the edge of the colony with a punch, and then a 3-point inoculation method (Torp and Langseth, Production of T-2 toxin by aFusarium resembling Fusarium poae. Mycopathologia, 1999, 147: 89-96) evenly inoculate the mycelial mass into T-2 toxin induction medium (recipe: yeast extract 20g, sucrose 150g, MgSO 4 ·7H 2 O 0.5g, 20g agar, add distilled water to make the volume to 1L, sterilize at 120°C for 20min before use) on the plate. After the parafilm was sealed, cultured in the dark at 25°C for 14 days. After the culture was ground and crushed by a stirrer, 5 g of the sample was weighed into a 50 ml Erlenmeyer flask, and then 25 ml of acetonitrile-water (volume ratio 84:16) was added, and the culture was shaken and extracted at 150 r / min for 16 hours....

Embodiment 3

[0057] Detection and identification of toxin produced by Fusarium culmorum

[0058] Fusarium culmorum (Table 2) strain mycelial total DNA extraction, specific PCR reaction system and amplification conditions are the same as the toxin detection and analysis steps of the strain Fusarium langsethiae in "Example 1". Chemical analysis showed that the strain could not produce T-2 toxin, and PCR amplification did not obtain any fragments. The results are completely consistent. The effect of the invention is as follows figure 2 The lane 12 is shown.

[0059]

[0060]

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Abstract

The invention belongs to the technical field of plant inspection and quarantine, and specifically relates to a molecular identification method of trichothecene type-A toxins of fusarium. The molecular identification method of the invention is characterized by comprising the steps of: designing a pair of specific primers through Tril13 gene information aiming at the trichothecene type-A toxins; separating the fusarium which generates type-A toxins of the fusarium to obtain specific segments (the length is 470 bp) of T-2 toxins, wherein the specific segments cannot be amplified in the fusarium which does not generate the type-A toxins of the fusarium; and adopting PCR (Polymerase Chain Reaction) reaction and utilizing a product from agarose gel electrophoresis to determine whether one fusarium can generate T-2 toxins directly according to existence and size of a product segment. The molecular identification method disclosed by the invention can be used for detecting trichothecene type-A toxins of fusarium in grains, feed and food safety.

Description

Technical field [0001] The invention belongs to the technical field of plant inspection and quarantine, and specifically relates to a molecular identification method of Fusarium trichothecenes A toxin. Background technique [0002] Fusarium head blight or scab (Fusarium head blight or scab) is an important disease that harms wheat, barley, corn and other crops worldwide. It not only seriously affects crop yields, but also Fusarium can produce a variety of toxic effects to humans and animals. The role of trichothecenes toxins reduces the quality of food or feed. Fusarium toxin has high stability and is not easily degraded. It can enter the food chain during food and feed processing, thereby threatening human and animal health. These toxins can inhibit the protein synthesis of eukaryotic cells and destroy the immune system of humans and animals. The poisoning is often accompanied by symptoms such as vomiting, diarrhea, and dizziness. Some toxins may even cause cancer. Trichothece...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/31C12N15/11C12Q1/68C12Q1/04
Inventor 廖玉才王建华李和平张静柏黄涛
Owner HUAZHONG AGRI UNIV
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