Primer and method for detecting fusarium graminearum with high resistance to carbendazim

A technology of Fusarium graminearum and carbendazim, applied in the field of molecular biology, can solve the problems of lack of fast and effective molecular detection methods, inability to distinguish mutation types of resistant strains, laborious and other problems, and achieve simple and rapid extraction methods, improve Specific, easily scalable effects

Active Publication Date: 2013-11-27
ZHEJIANG UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0005] Traditional bioassays for drug resistance are time-consuming, laborious and unable to differentiate mutation types in resistant strains
At present, detection techniques such as ASO-PCR, PIRA-PCR and Tetra-primer ARMS-PCR have been reported for point mutations F167Y, F200Y, E198Q and E198K (such as the Chinese patent document "Fusarium graminearum identification and its One-tube detection method for confirmation of moderate drug resistance to carbendazim"; Chinese patent document with publication number CN101985653A "Molecular detection method for strains with intermediate drug resistance of Fusarium graminearum to carbendazim"), but for The point mutation E198L that causes high levels of carbendazim resistance lacks rapid and effective molecular detection methods

Method used

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  • Primer and method for detecting fusarium graminearum with high resistance to carbendazim
  • Primer and method for detecting fusarium graminearum with high resistance to carbendazim
  • Primer and method for detecting fusarium graminearum with high resistance to carbendazim

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Embodiment 1

[0038] 1. Strain type

[0039] F. avenaceum, F. acuminatum, F. semitectum, F. subglutinans, F. culmorum 1 strain each, and 11 strains of F. graminearum. The 11 Fusarium graminearum strains included 2 carbendazim-susceptible strains (PH-1, HN9), 2 low-antibacterial strains containing the E198Q point mutation (JM51, YL10), and 2 medium-antibacterial strains containing the F167Y point mutation ( YU59, BM10), 2 medium antibacterial strains containing F200Y point mutation (BY74, QJ42), 1 medium antibacterial strain JL45 containing E198K point mutation, and 2 high antibacterial strains containing E198L point mutation (JM2, JM5).

[0040] The above-mentioned strains are all common plant pathogenic fungi, which are preserved in the Institute of Biotechnology of Zhejiang University, and can also be obtained by conventional methods of separation and purification of fungi.

[0041] 2. Primer synthesis

[0042] The study found that the high level of resistance of Fusarium graminearum t...

Embodiment 2

[0061] Example 2 Detection of Fusarium graminearum with high carbendazim resistance in ascus shells on rice piles in different regions

[0062] In April 2012, rice stumps with asthecia were collected from wheat fields in Hai'an and Jingjiang, Jiangsu, and Fanchang and Chizhou, Anhui (approximately 80 asthecias per field).

[0063] Place the ascothecia collected on rice stumps into a 1.5 mL Eppendorf tube and add 100 μl of DNA extraction buffer (1-2% polyvinylpyrrolidone (PVP), 200 mM Tris-HCl, 50 mM EDTA, 200 mM NaCl, 1% SDS, with ddH 2 O is the solvent, pH 8.0), and grind for 1 min with a pointed glass rod mounted on a commonly used electrician’s electric hand, then add 400 μl of DNA extraction buffer (DEB, DNA extraction buffer) to the centrifuge tube, vortex and mix , stand at room temperature for 10 minutes; centrifuge the above mixture at 4°C and 15,000 rpm for 5 minutes, then transfer the supernatant to another centrifuge tube, then add 750 μl of absolute ethanol, and mi...

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Abstract

The invention discloses primers and a method for detecting fusarium graminearum with high resistance to carbendazim. The base sequences of the primers are as follows: the base sequence of the forward primer is 5'-TTG CAA TTG CAA ATT TCA ACG TG-3'; the base sequence of the reverse primer is 5'-CGT TAT CGA TAC AGA AGG TAA-3', or 5'-CGT TAT CGA TAC AGA AGG TTA-3'. The invention provides two groups of primers, PCR augmentation is performed through any one of the primers so as to detect the fusarium graminearum which has high resistance to carbendazim, and the specificities of the primers are high; in addition, the method disclosed by the invention can quickly and accurately detect the fusarium graminearum with the high resistance to carbendazim, the whole process only costs about two hours, and the operation is fast and accurate.

Description

technical field [0001] The invention relates to the field of molecular biology, in particular to a primer and a method for detecting highly carbendazim-resistant Fusarium graminearum strains. Background technique [0002] Wheat head blight caused by Fusarium graminearum species complex seriously threatens the safety of wheat production. In addition, Gibberella also produces trichothecenes, which seriously affect food safety. [0003] In my country, in recent years, due to the change of straw returning to the field and climatic factors, the incidence of head blight has been aggravated. Due to the lack of germplasm materials with high levels of resistance, chemical control is currently the main method used in production to control wheat head blight. Carbendazim fungicide is the main agent for the control of wheat head blight, but due to the long-term and extensive use of this type of agent, Fusarium graminearum has carbendazim resistance in Zhejiang, Jiangsu and Anhui and ot...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/11C12Q1/68C12Q1/04
Inventor 尹燕妮刘晔马忠华
Owner ZHEJIANG UNIV
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