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Method for screening anoplophora glabripennis attracting components and preparing anoplophora glabripennis attractant

A glaucous longhorn beetle and attractant technology, applied in the field of forestry and forest protection, can solve problems such as the difficulty of screening out the attracting active ingredients, single host plants or herbivorous insects

Inactive Publication Date: 2017-09-15
HEILONGJIANG PROV FOREST PROTECTION INST
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The purpose of the present invention is to solve the shortcomings of the prior art from the perspective of a single host plant or herbivorous insect, and it is difficult to screen out efficient attractant active ingredients, and propose a method for screening efficient attractant ingredients and preparing an attractant for Beetle glabrata Methods

Method used

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  • Method for screening anoplophora glabripennis attracting components and preparing anoplophora glabripennis attractant
  • Method for screening anoplophora glabripennis attracting components and preparing anoplophora glabripennis attractant
  • Method for screening anoplophora glabripennis attracting components and preparing anoplophora glabripennis attractant

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specific Embodiment approach 1

[0027] Specific embodiment one: a kind of method of screening Beetleus glabra's attractant composition comprises the following steps:

[0028] Step 1: extracting and purifying the total RNA of the antennal sensilla of Beetleus glabrata;

[0029] Step 2: Obtain the full length of the odor-binding protein of Beetle glabrata;

[0030] Step 3: Construct the prokaryotic expression vector of the odorant binding protein of Beetle glabrata;

[0031] Step 4: Induction and purification of the recombinant odorant-binding protein of Beetle glabrata;

[0032] Step 5: Screening and identification of the attractant components of the bald-shouldered beetle.

specific Embodiment approach 2

[0033] Specific embodiment two: the difference between this embodiment and specific embodiment one is: the specific process of the extraction and purification of the total RNA of the antennal sensilla in the step one is as follows:

[0034] Use Trizol reagent to extract total RNA from the antennae of Beetleus glabrata, use non-enzymatic DNA removal-A (DNA-Be-Gone-A) to remove residual DNA in the total RNA, and use 0.01 agarose gel electrophoresis to detect the integrity of the total RNA; use SMART IVoligonucleotide Reverse transcription with CDSⅢ / 3'PCR primer (primer), and incubate at 42°C for 1 hour to generate the first strand of cDNA.

[0035] Other steps and parameters are the same as those in Embodiment 1.

specific Embodiment approach 3

[0036] Specific embodiment three: the difference between this embodiment and specific embodiment one or two is that: the specific process of obtaining the full length of the odor-binding protein of Beetleus glabrata in said step two is:

[0037] Primer design for Agla-OBP1, Agla-OBP2,

[0038] Agla-OBP1 primer sequence:

[0039] Forward: 5'-ATGAAACTTTTTGTATTCGTCC-3'

[0040] Reverse: 5'-TTAGGGCAAGAAATAATTCGAG-3'

[0041] Agla-OBP2 primer sequence:

[0042] Forward: 5'-ATGTCGCTCAGAATCGTTATCG-3'

[0043] Reverse: 5'-CTATACCAGGAACCAGTTCTCC-3'

[0044] Amplification, cloning and sequencing of the cDNA of odorant-binding proteins Agla-OBP1 and Agla-OBP2 from Beetle glabrata. Use the first strand of cDNA synthesized by reverse transcriptase as a template, and use primers to amplify it by PCR; reaction system: Pfu 0.5 μl, PfuBuffer+Mg 2+ (SO 4 2- ) 5.0 μl, dNTP 4.0 μl, cDNA 1.0 μl, Agla-OBP1-forward 1.0 μl, Agla-OBP1-reverse 1.0 μl, ddH 2 O 37.5μl; PCR amplification program:...

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Abstract

The invention relates to a method for screening anoplophora glabripennis attracting components and preparing an anoplophora glabripennis attractant. According to the method, the disadvantages that the methods in the prior art are disclosed from host plants or phytophagous insects in one aspect, and therefore, efficient attracting active components are difficult to be screened are solved. The method comprises the following steps: (1) extracting and purifying total RNA of anoplophora glabripennis antennal sensilla; (2) acquiring the overall length of odorant binding proteins of the anoplophora glabripennis; (3) constructing prokaryotic expression vectors of the odorant binding proteins of the anoplophora glabripennis; (4) inducing and purifying the odorant binding proteins of the anoplophora glabripennis; and (5) screening the anoplophora glabripennis attracting components, and determining an optimal formula of the anoplophora glabripennis attractant by virtue of GC-EAG, indoor smell tropism determination and field trapping experiments. The method is applied to the technical field of forestry forest protection science.

Description

technical field [0001] The invention relates to the technical field of forestry and forest protection. Background technique [0002] The prior art confirms that along with the maturity of chemical, ecological and molecular biotechnology and research methods, the comprehensive insect management technology has also undergone a qualitative leap. This leap is reflected in the comprehensive utilization of chemical communication between insects and host plants, and between species. Therefore, research on insect control measures is carried out from the fields of ecology, chemistry and biotechnology, that is, the use of host insects, host plants, natural enemies, etc. can release specific volatile substances that attract, repel or not attract host insects to host plant selection. Sexual chemicals to achieve the purpose of insect prevention. [0003] Beetle glabrata is a stem-boring omnivorous insect, but the host plants have certain differences in the attraction and damage to the b...

Claims

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Application Information

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IPC IPC(8): C12N15/10C12N15/70C12N15/66C07K14/435C07K1/22G01N33/68
CPCC07K14/43563C12N15/10C12N15/66G01N33/68G01N2333/43552
Inventor 马晓乾范海娟祖榕逄弘扬赵红盈冯磊孙强
Owner HEILONGJIANG PROV FOREST PROTECTION INST
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