Detection primers and molecule detection method for pathogenic fungi of austral puccinia sorghi

A technology for corn rust and pathogenic fungi is applied in the field of molecular biology to achieve the effects of low test cost, simple and fast operation, and good practicability

Inactive Publication Date: 2011-09-07
HENAN AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the research on molecular detection of corn rust fungus has not been reported at home and abroad.

Method used

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  • Detection primers and molecule detection method for pathogenic fungi of austral puccinia sorghi
  • Detection primers and molecule detection method for pathogenic fungi of austral puccinia sorghi
  • Detection primers and molecule detection method for pathogenic fungi of austral puccinia sorghi

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] Example 1 Field corn rust monitoring comparison test

[0027] (1) Sample collection

[0028] From September 1 to September 9, 2008, 20 maize leaf samples were randomly collected in Zhoukou Taikang, Zhumadian Xiping, Zhengzhou Maozhuang, Kaifeng Lankao, Xinxiang Yuanyang, Nanyang Fangcheng, and Hebi Jun counties respectively. Store at -80°C for later use.

[0029] (2) Synthesize primers for detection of southern type rust according to the following nucleotide sequence (the same below):

[0030] Upstream primer: 5'- CTC CAA GAA CTT CCT CCT C -3',

[0031] Downstream primer: 5'- TGA CAT GAA GTA GAA ATT CT -3'.

[0032] (3) Detection of corn rust

[0033] ① Extraction of genomic DNA from maize leaves;

[0034] ②PCR amplification, 25μL reaction system was placed in the PCR thin-walled tube in turn:

[0035] 10′Buffer 2.5μL, 25mmol / LMgCL 2 2.0μL, 2.5mmol / LdNTP 2.0μL, 2.5μL each of the above-mentioned southern rust fungus upstream primer and 2mmol / L downstream primer, ...

Embodiment 2

[0039] Example 2 Comparison experiment of field corn rust monitoring

[0040] (1) Sample collection

[0041] On February 17, 2009, Chongpo Town, Sanya City, Hainan Province, August 26, Changchun City, Jilin Province, and September 24, Qianjiang Xinhua City, Chongqing City, respectively randomly took 10, 7, and 4 corn leaf samples,- Store at 80°C for later use.

[0042] (2) Detection of corn rust

[0043] ① Extract genomic DNA from maize leaves;

[0044] ②PCR amplification, 25μL reaction system was placed in the PCR thin-walled tube in turn:

[0045] 10′Buffer 2.5μL, 25mmol / LMgCL 22.0μL, 2.5mmol / LdNTP 2.0μL, 2.5μL each of the upstream primer and downstream primer 2mmol / L of the above-mentioned southern rust fungus of corn, 10~50ng of corn leaf genomic DNA, 1.0U of Taq DNA polymerase, and finally with sterile deionized Make up to 25 μL with water; centrifuge for 10 sec, put the PCR thin-walled tube into the PCR machine for amplification; the amplification conditions are: fi...

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Abstract

The invention relates to specific primers and a molecule detection method for pathogenic fungi of austral puccinia sorghi. The specific primers comprise a forward primer 5'-CTCCAAGAACTTCCTCCTC-3' and a reverse primer 5'-TGACATGAAGTAGAAATTCT-3'. The primers are subjected to PCR (Polymerase Chain Reaction) amplification and agarose gel electrophoresis, and the bacteria bearing condition of corns infield can be detected by adopting the primers and amplification products with segment length of 540bp, obtained through PCR amplification, wherein PCR amplification comprises the following steps of: pre-modifying for 5 minutes at 95 DEG C; then modifying for 45 seconds at 94 DEG C, annealing for 45 seconds at 55 DEG C, stretching for 90 seconds at 72 DEG C, and repeating the steps for 30 cycles; and finally stretching for 10 minutes at 72 DEG C. The primers disclosed by the invention have strong specificity for corn puccinia strilformis, can be used for detecting corn rust diseases and has high sensitivity up to 0.001 ng/mu m L. The detecting method disclosed by the invention has good practicability and is simple, convenient and quick to operate.

Description

technical field [0001] The invention relates to the field of molecular biology, in particular to a specific primer and a molecular detection method for the pathogenic fungus of southern corn rust. Background technique [0002] Corn rust is an increasingly serious fungal disease of corn worldwide. There are three kinds of corn rusts that have been reported so far, namely, the common corn rust caused by Puccinia sorghhi Schw., the southern corn rust caused by Puccinia polysora Underw. and the corn rust caused by corn hulls. Tropical corn rust caused by the fungus [Physopella zeae(Mains) Cummins & Ramachar]. [0003] Corn rust occurs in a wide range in my country, covering the main corn producing areas in the north and south. There are two types of common rust and southern rust. Common rust mostly occurs in the northeast and northwest corn areas with low temperature and high humidity; southern rust occurs in Taiwan , Hainan, and Yuanjiang, Yunnan first occurred. In recent yea...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68
Inventor 郑文明邢国珍蒋士君许君袁红霞王明凤杨文博张磊
Owner HENAN AGRICULTURAL UNIVERSITY
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