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Tea geometrid odorant binding protein-based plant attractant screening method

An odor-binding protein and screening method technology, applied in the field of screening plant-derived attractants, can solve problems such as difficulty in effective design and development, complex composition of volatiles in tea leaves, etc.

Inactive Publication Date: 2013-11-27
HANGZHOU ACAD OF AGRI SCI +1
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Among them, using its olfactory behavior to design its attractant has become an important means of comprehensive management of tea geometrids, and the main representative technology is to identify the volatiles of tea geometrids on tea tree leaves by means of chemical instrument analysis. The composition of the compound is very complex, so it is difficult to effectively design and develop an olfactory attractant formula suitable for the tea geometrid

Method used

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  • Tea geometrid odorant binding protein-based plant attractant screening method
  • Tea geometrid odorant binding protein-based plant attractant screening method
  • Tea geometrid odorant binding protein-based plant attractant screening method

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Effect test

Embodiment 1

[0024] 1) The specific steps of collecting total RNA from the antennae of the tea looper;

[0025] Material: Tea looper catches larvae for Hangzhou tea gardens, and artificially raises them in the laboratory until they become adults for experimental use. The larva feeding conditions are: temperature 26±1℃, L:D=12:12, RH 70%~80%. The TRIzol method was used to extract the total RNA of the antennae of the adult tea looper, and the Super-Script? Ⅱ Reverse Transcriptase System (Takara) was used to reverse transcription to obtain the first strand of cDNA.

[0026] 2) The specific steps of obtaining the full length of the odor binding protein of tea looper by RT-PCR;

[0027] ①The primer design of EoblGOBP2. According to the gene sequence of the common odor-binding protein GOBP2 (EoblGOBP2) in Genbank (Genbank accession number: ACN29681.1 (12), Primer Premier 5.0 software was used to design primers. In order to facilitate the later expression of the target gene cloned into the expression ...

Embodiment 2

[0052] Test to prove the effectiveness of the plant-derived attractant obtained by the present invention.

[0053] 1) Test insects. The mature larvae of tea looper were collected from the artificially raised population in this laboratory. The feeding conditions are: temperature (26±1) ℃, relative humidity (75±5)%, photoperiod L: D = 12: 12 (light 12 h, dark 12 h). After pupation and emergence, the adults at the sexual maturity stage were collected for experiment, and they were starved for 3 h before the experiment.

[0054] 2) Experimental instruments and reagents. The experimental instrument was an antenna potentiometer (Syntech, Netherlands); various tea tree leaf volatile standard products were purchased from Bai Lingwei, and each compound was dissolved in liquid paraffin and prepared into a 0.1 (v / v) solution for EAG determination. Drop 10 uL of the sample solution to be tested on the filter paper strip (2 cm×0.5 cm) in advance, let the solvent volatilize for a while, then p...

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Abstract

The invention discloses a tea geometrid odorant binding protein-based plant attractant screening method and belongs to the technical field of bioengineering. The tea geometrid odorant binding protein-based plant attractant screening method comprises the following processes of collecting tea geometrid antenna total RNA, acquiring total length of a tea geometrid odorant binding protein by RT-PCR, constructing a prokaryotic expression vector of the tea geometrid odorant binding protein, inducing the expression of the recombinant tea geometrid odorant binding protein by IPTG, carrying out the purification of the recombinant tea geometrid odorant binding protein by nickel sepharose gel affinity column, acquiring a binding reaction spectrum of the recombinant tea geometrid odorant binding protein and tea leaf smell volatiles by a competitive fluorescent combination method, and determining that the tea leaf smell volatile reducing relative fluorescence intensity of 1-NPN to less than 50% and having the binding constant of 13-45 micromoles per liter is the plant attractant of tea geometrid. The tea geometrid odorant binding protein-based plant attractant screening method provides a novel means for screening and designing a formula of a plant tea geometrid smell information attractant.

Description

Technical field [0001] The invention belongs to the technical field of bioengineering, and specifically relates to a method for screening a plant-derived attractant based on the tea looper odor binding protein. Background technique [0002] Tea looper Ectropis obliqua Prout, belonging to Lepidoptera, Geometridae, is a major tea pest widely distributed in various tea-producing provinces in the Yangtze River Basin. Because it likes to eat tea leaves, it has a large number of generations and reproduces quickly. When it occurs, all leaves are eaten. Light, causing the tea industry to reduce production or even fail. For a long time, the control of tea looper mainly relied on chemical pesticide control, which easily caused the problem of pesticide residues in commercial tea, which seriously threatened the normal development of my country’s tea trade industry. In view of this, it is necessary to develop a new type of pollution-free tea in addition to chemical pesticides. Inchworm biolo...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N21/64C12N15/12C12N15/63C07K14/435
Inventor 余继忠李红亮赵磊黄海涛崔宏春
Owner HANGZHOU ACAD OF AGRI SCI
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