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Application of specific primer for identifying physiological race THTS of puccinia triticcina

A technology for wheat leaf rust and physiological races, which is applied in the application field of identifying the physiological race of wheat leaf rust THTS specific primers, and can solve the problems of few molecular markers and inability to identify races with DNA molecular markers.

Inactive Publication Date: 2015-10-14
HEBEI AGRICULTURAL UNIV.
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  • Abstract
  • Description
  • Claims
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Problems solved by technology

Among them, SSR markers have been widely used because of their high polymorphism, co-dominance, and good stability. However, at present, there are few molecular markers specific to physiological races of P. tritici. species identification

Method used

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  • Application of specific primer for identifying physiological race THTS of puccinia triticcina

Examples

Experimental program
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Effect test

experiment example

[0025] Obtaining of Physiological race THTS and EST-SSR marker Ptssr0125 of Phytophthora triticifolia:

[0026] 21 pairs of EST-SSR primers were selected, and all primer sequences were derived from Wang et al. The EST-SSR primers for wheat leaf rust developed in 2010, use the genomic DNA of 213 wheat leaf rust strains as templates for PCR detection, the 20μL PCR reaction system is: template DNA 60 ng, Taq enzyme 0.5 U, upstream and downstream Primers (5 μmol L -1 ) each 0.4 μL, dNTP (10 mmol·L -1 ) 0.4 μL, 10 × PCR buffer 2 μL, supplement the reaction system with sterile ultrapure water to 20 μL. The PCR reaction program was as follows: first at 94°C for 5 min; then at 94°C for 30 s, at 55°C for 30 s, and at 72°C for 1 min, a total of 35 cycles; finally at 72°C for 5 min, and stored at 4°C.

[0027] Then carry out 10% (w / v) non-denaturing polyacrylamide gel electrophoresis detection on the PCR amplification product according to the following method: take 20 μL of the ampl...

Embodiment 1

[0030] 1. Extract the genomic DNA of 213 leaf rust strains as templates;

[0031] 2. Using Ptssr0125 as a primer and 213 genomic DNAs of leaf rust as a template, carry out PCR amplification. The 20 μl PCR reaction system is: template DNA 60 ng, Taq enzyme 0.5 U, downstream primer (5 μmol L -1 ) each 0.4 μL, dNTP (10 mmol·L -1 ) 0.4 μL, 10 × PCR buffer 2 μL, supplement the reaction system with sterile ultrapure water to 20 μL. The reaction program was: first at 94 °C for 5 min; then 35 cycles: 94 °C for 30 s, 55 °C for 30 s, 72 °C for 1 min; finally 72 °C for 5 min, and stored at 4 °C.

[0032] 3. Separation and detection of the amplified products: add non-denaturing loading indicator to the amplified products, and then separate them by electrophoresis in a denatured polyacrylamide gel with a concentration of 10% (w / v), and develop the color by silver staining. In the PCR amplification pattern detected by polyacrylamide gel electrophoresis, the bands with 200bp and 178bp at ...

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Abstract

The invention discloses application of a primer for identifying physiological race THTS of puccinia triticcina and a THTS race-specific DNA (deoxyribonucleic acid) sequence. The primer sequence is a primer pair which is composed of forward and reverse nucleotide sequences of primer Ptssr0125 in a sequence table. Fast physiological race identification can be carried out on the puccinia triticcina with an EST-SSR (expressed sequence tag-simple sequence repeat) molecular marker.

Description

technical field [0001] The invention relates to a primer sequence for identifying physiological races of wheat leaf rust and its application. Background technique [0002] by wheat leaf rust ( Puccinia triticina ) caused by wheat leaf rust is one of the common diseases in wheat regions in the world. Its occurrence and prevalence seriously affect the yield and quality of wheat, and it is an important obstacle affecting the world's wheat production. Depending on the wheat variety and the disease onset period, it can cause 7%-30% or even more than 50% yield loss (Huerta-Espino et al. 2011). Practice has proved that the most economical, safe and effective way to control the disease is the application of disease-resistant varieties. However, the loss of rust resistance of cultivars is a serious problem in the application of disease-resistant cultivars, mainly due to the variation of leaf rust fungi and the emergence of new virulent races. Therefore, monitoring and epidemic for...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12Q1/04C12N15/11C12R1/645
Inventor 闫红飞张林亚杨文香刘大群
Owner HEBEI AGRICULTURAL UNIV.
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