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Specific primer pair for detecting anti-ALS inhibitor herbicide brome, kit and detection method and application

An inhibitor and herbicide technology, applied in the field of molecular biology, can solve the problems of inability to directly guide scientific drug use, occupy a lot of space, and have a long detection period, and achieve the effects of low cost, simple operation and high accuracy.

Pending Publication Date: 2022-01-18
INST OF CEREAL & OIL CROPS HEBEI ACAD OF AGRI & FORESTRY SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

This method can intuitively evaluate the resistance level of weeds to herbicides, but it takes up a lot of space and workload, the detection cycle is long, and it cannot directly guide scientific medication

Method used

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  • Specific primer pair for detecting anti-ALS inhibitor herbicide brome, kit and detection method and application
  • Specific primer pair for detecting anti-ALS inhibitor herbicide brome, kit and detection method and application
  • Specific primer pair for detecting anti-ALS inhibitor herbicide brome, kit and detection method and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0043] 1. Extraction of Brome Genomic DNA

[0044] Take 0.2 g of brome seedling leaves, add liquid nitrogen into the mortar and grind thoroughly, and use the CTAB method to extract brome genomic DNA, and the DNA concentration is greater than 50 ng / μL.

[0045] 2. Primer synthesis

[0046] The primers shown in SEQ ID NO.1 and SEQ ID NO.2 were synthesized by General Biosystems (Anhui) Co., Ltd. Different brome populations were used for primer PCR amplification, and the results showed that the target band was of correct length, clear and bright, indicating that the primer amplification efficiency was high and the specificity was strong ( figure 1 ).

[0047] 3. PCR reaction system for detection of brome ALS mutation site

[0048] PCR reaction system, including 0.5 μL of upstream and downstream primers (concentration 10 pmol / L), 12.5 μL of 2×Es TaqMasterMix, 1 μL of DNA template, and 10.5 μL of double distilled water.

[0049] 4. Reaction program for amplifying brome ALS gene ...

Embodiment 2

[0054] The brome population suspected of developing resistance was collected, and about 0.2 g of fresh brome leaves were quickly added to liquid nitrogen for grinding, and the genomic DNA was extracted by the CTAB method, and the DNA concentration was greater than 50 ng / μL. See Example 1 for the test method. The specific primers amplified the brome ALS gene with good specificity, and the amplified fragment of the ALS gene coding region was about 1700bp ( image 3 ). The primers were used to amplify the resistant and sensitive brome ALS genes, the PCR products were sequenced, and the mutation sites were found by sequence comparison analysis ( Figure 4 , Figure 5 ).

[0055] 18 brome populations collected from different winter wheat fields in Hebei Province were detected according to the detection method of the present invention. The results showed that the 197th proline of ALS in 11 populations was mutated to serine, and the ALS 197th position in 3 populations was mutated ...

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Abstract

The invention provides a specific primer pair for detecting anti-ALS inhibitor herbicide brome, a kit, a detection method and application, and relates to the technical field of molecular biology. A specific strip is good in specificity, and a PCR target strip is clear. By utilizing the specific primer pair or the kit containing the specific primer pair, anti-ALS inhibitor herbicide bromegrass can be accurately and specifically identified, and the detection method is simple and convenient to operate, high in speed, low in cost and high in accuracy. The invention further provides a detection method based on the specific primer pair and the kit, ALS mutation sites and amino acid mutation types of field suspected resistant brome can be efficiently and accurately detected, and whether ALS inhibitor herbicides can be continuously used for preventing and removing resistant brome or not can be guided.

Description

technical field [0001] The invention belongs to the technical field of molecular biology, and in particular relates to a pair of specific primers, a kit, a detection method and an application for detecting the anti-ALS inhibitor herbicide brome. Background technique [0002] Wheat is one of the main food crops in my country, and its yield is closely related to the national economy and people's livelihood. Weed damage is one of the main factors leading to the reduction of wheat yield. With the change of global environment, the change of agricultural planting system, the application of mechanized agricultural operation and the introduction of wheat in different regions, the population of weeds in wheat fields has changed. Currently brome has been succeeded as one of the vicious weeds in wheat field. On average, brome-occurring plots cause a 15-30% reduction in wheat yield, and a severe plot has a wheat yield loss of about 70%. At present, flusasulfuron-methyl is mainly used...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6895C12Q1/6858C12N15/11
CPCC12Q1/6895C12Q1/6858C12Q2600/13C12Q2600/156C12Q2600/106C12Q2531/113C12Q2565/125
Inventor 许贤王贵启刘小民李秉华赵铂锤申贝贝王建平崔海燕祁志尊
Owner INST OF CEREAL & OIL CROPS HEBEI ACAD OF AGRI & FORESTRY SCI
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