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Molecular identification method of trichothecene type-A toxins of fusarium

A technology of trichothecenes and Fusarium, applied in the fields of botany equipment and methods, biochemical equipment and methods, microbial determination/inspection, etc., can solve the problem that the results cannot be consistent with each other, and the toxin-producing species have not undergone chemical analysis Validation, low credibility, etc.

Inactive Publication Date: 2012-05-09
HUAZHONG AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

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 bacterial strain Fusarium langsethiae (see Table 1) was activated and cultivated on the PDA plate for 5 days, a mycelial block with a diameter of 2 cm was obtained at the edge of the colony with a puncher, 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 mycelium block into T-2 toxin induction medium (formula: yeast extract 20g, sucrose 150g, MgSO 4 ·7H 2O 0.5g, 20g agar, add distilled water to make up to 1L, use after sterilizing at 120°C for 20min) on the plate. After sealing with parafilm, culture at 25°C in the dark for 14 days. After the culture was pulverized 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 extracted by shaking at 150 r / min for 16 h. ...

Embodiment 2

[0052] Detection and Identification of Toxin Produced by Fusarium poae

[0053] After bacterial strain Fusariumpoae (see Table 1) was activated and cultivated on the PDA plate for 5 days, a 2 cm mycelium piece was obtained with a puncher at the edge of the colony, 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 mycelium block to T-2 toxin induction medium (formula: yeast extract 20g, sucrose 150g, MgSO 4 ·7H 2 O 0.5g, 20g agar, add distilled water to make up to 1L, use after sterilizing at 120°C for 20min) on the plate. After sealing with parafilm, culture at 25°C in the dark for 14 days. After the culture was ground and pulverized by a stirrer, 5 g of the sample was weighed into a 50 ml Erlenmeyer flask, and then 25 ml of acetonitrile-water (84:16 by volume) was added, and extracted by shaking at 150 r / min for 16 h. The extract was filtered through ...

Embodiment 3

[0057] Detection and Identification of Toxin Produced by Fusarium culmorum

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

[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 in particular relates to a molecular identification method of fusarium trichothecene family A toxoids. Background technique [0002] Fusarium head blight (Fusarium head blight or scab) caused by Fusarium is an important disease that harms wheat, barley, corn and other crops worldwide. The active trichothecenes can reduce the quality of food or feed. Fusarium toxin has high stability and is not easy to degrade. It can enter the food chain along with food and feed processing, and then threaten the health of humans and animals. These toxins can inhibit the protein synthesis of eukaryotic cells and destroy the immune system of humans and animals. After poisoning, they are often accompanied by symptoms such as vomiting, diarrhea, and dizziness. Some toxins may even cause cancer. Trichothecenes are the most important Fusarium toxins, mainly including A and B. T-2 toxin be...

Claims

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Application Information

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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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