Molecular detection method of resistance of diamond back moth against avermectin target

A technology for the detection of abamectin targets and molecules, applied in the biological field, can solve the problems of long cycle time, low sensitivity, and high material requirements, and achieve the effect of high sensitivity, low material requirements, and accurate detection

Inactive Publication Date: 2016-06-29
NANJING AGRICULTURAL UNIVERSITY
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  • Application Information

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Problems solved by technology

[0005] The purpose of the present invention is to solve problems such as low sensitivity, long period, and high material requirements of the detection method of diamondback moth in the prior art for avermectin resistance. Rapid and Accurate Molecular Detection of Abamectin Resistance Target Mutations in Plutella xylostella

Method used

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  • Molecular detection method of resistance of diamond back moth against avermectin target
  • Molecular detection method of resistance of diamond back moth against avermectin target
  • Molecular detection method of resistance of diamond back moth against avermectin target

Examples

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Effect test

Embodiment 1

[0028] In this example, three groups of Plutella xylostella were selected for bioassay, and the ROTH strain was a control strain sensitive to insecticides that had been raised in the laboratory for many years. In this example, the lethal concentration of the material to abamectin was used as the baseline to measure other species. The resistance level of the strain to abamectin; the ROTH-Abm strain is a near isogenic line material resistant to abamectin, and its resistance level to abamectin is 10100 times; the Abm-Unsel strain is ROTH- The resistance of Abm test insects was significantly reduced to 1695 times when they were raised for more than one year without contact with any pesticide. The relevant biometric data are as follows:

[0029]

[0030] According to the preferred analysis and detection technology of the present invention, its specific implementation steps include:

[0031] 1. Randomly select 17, 19 and 26 fourth-instar larvae from ROTH, ROTH-Abm and Abm-Unsel ...

Embodiment 2

[0046] This example illustrates the preferred technical solution of the present invention for detecting the detection method of the PxGluCl gene mutation gene frequency at position 309 carried by the Abm-Unsel strain of diamondback moth Abm-Unsel under different drug treatment concentrations.

[0047] 1. Extraction of genomic DNA from single Plutella xylostella larvae: Randomly pick the 3rd instar larvae of Plutella xylostella of the Abm-Unsel strain, and treat a batch of test worms with 80ppm, 20ppm and 0ppm abamectin respectively, and check the results after 48 hours, it is normal Surviving larvae using AXYprep TM The MultisourceGenomicDNA MiniprepKit Genome Kit extracts whole genome DNA.

[0048] 2. PCR amplification of the target fragment of the glutamic acid chloride ion channel gene was carried out with the gene DNA templates (3 groups) of the surviving diamondback moth after being treated with various concentrations of abamectin.

[0049] (1) Design specific primers fo...

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Abstract

The present invention discloses a molecular detection method of resistance of diamond back moth against an avermectin target. According to the method, a specific primer designed according a diamond back moth glutamate chloride ion channel gene sequence is used, extracted diamond back moth single-head genomic DNA is used as a template for PCR amplification of glutamate chloride ion channel gene fragments, diamond back moth individual genotypes (sensitive homozygote SS, resistant heterozygote RS and resistant homozygote RR) can be distinguished by analysis of a target fragment purified product direct sequencing chromatogram, and population sample resistance allele mutation frequency can be detected. A detection method of diamond back moth glutamate chloride ion channel gene A309V mutation and a special primer sequence are disclosed, and the method has the advantages of being simple, rapid and high in accuracy, can be used to monitor the occurrence and development trend of allele frequency of the resistance of a diamond back moth field population against avermectin and drug resistance, and provides an important basis for adjustment of diamond back moth chemical control strategies and formulation of measures to delay the development of resistance.

Description

technical field [0001] The invention relates to a molecular detection method for the resistance of diamondback moth to avermectin target, belongs to the field of biotechnology, and is specially used for the target resistance of diamondback moth to the broad-spectrum insecticide and acaricide avermectin (composed of AvermectinB1a and AvermectinB1b mixed) Highly sensitive and rapid molecular detection. technical background [0002] According to statistics, in recent years, my country's vegetable planting area is close to 300 million mu per year, and it has become the world's largest planter and producer of cruciferous vegetables. The reduction in vegetable production and management costs caused by diamondback moth damage is extremely high. Plutella xylostella belongs to Lepidoptera Plutellaidae and is an important pest of cruciferous vegetables worldwide, causing economic losses of 4 to 5 billion US dollars every year. There are more than 40 species of host plants in this spe...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12N15/11
Inventor 王兴亮吴益东杨亦桦武淑文
Owner NANJING AGRICULTURAL UNIVERSITY
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