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Imidaclothiz antibody specifically-bound phage display polypeptide and use thereof

A chlorothialine and specificity technology, which is applied in the application field of detection of chlorothialine, can solve problems such as unseen peptides, and achieve the effects of high accuracy, high sensitivity, and strong specificity

Inactive Publication Date: 2017-05-31
NANJING AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

But so far, there have been no researches and reports on peptides specifically binding to chlorothialine antibodies and their applications.

Method used

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  • Imidaclothiz antibody specifically-bound phage display polypeptide and use thereof
  • Imidaclothiz antibody specifically-bound phage display polypeptide and use thereof
  • Imidaclothiz antibody specifically-bound phage display polypeptide and use thereof

Examples

Experimental program
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Effect test

Embodiment 1

[0028] Example 1 Panning and preparation of chlorothialine antibody specific binding polypeptide

[0029] 1. Panning of phage clones that specifically bind to chlorothialine antibody

[0030] According to the cycle of "adsorption-washing-elution-amplification", after three rounds of panning, the specific operation is as follows:

[0031] (1) Take 100 μL of 75 μg / mL protein A column-purified chlorothialine antibody and add it to a microtiter plate, and coat at 4°C overnight, with a total of three wells;

[0032] (2) Take a small amount of Escherichia coli ER2738 and spread it on the LB+Tet plate, culture overnight at 37°C;

[0033] (3) Wash the ELISA plate in step (1) 5 times with PBST, add 300 μL of 5% milk, incubate at 37°C for 2 hours, wash 5 times with PBST, store at 4°C for later use;

[0034] (4) Add 100 μL 2×10 11 phage (containing 3% milk), shake slightly at room temperature for 1 hour;

[0035] (5) Take 20mL of LB medium and add it to a 250mL Erlenmeyer flask, add ...

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Abstract

The invention belongs to the technical field of biology and relates to an imidaclothiz antibody specifically-bound polypeptide and use of the polypeptide in detection of imidaclothiz. According to the use, screening of three times is carried out on a random cyclo-heptapeptide library, a cyclo-octopeptide library and a linear-dodecapeptide library for phage display by using a protein A column purified imidaclothiz antibody by adopting a phage display technology, so as to screen out phage clones bound to the imidaclothiz antibody; and amplification and plasmid extraction are carried out on a plurality of randomly-selected phage clones, positive phage clones are selected by phage ELISA (Enzyme-Linked Immunosorbent Assay), and sequencing is carried out on the positive clones so as to obtain a polypeptide sequence. The invention further relates to the use of the phage display polypeptide in the detection of the imidaclothiz. The phage ELISA established by using the phage display polypeptide can be used for detecting imidaclothiz residues in surroundings and agricultural products in a rapid, sensitive, simple, convenient and cheap manner.

Description

technical field [0001] The invention belongs to the field of biotechnology, and relates to a polypeptide with specific binding to chlorothialine antibody, including the application of the polypeptide in detecting chlorothialine. Background technique [0002] Chlorothialine (imidaclothiz) is a new neonicotinoid insecticide developed by Nantong Jiangshan Pesticide Chemical Co., Ltd. It can be used to control sucking mouthpart pests, such as aphids, leafhoppers, planthoppers, thrips, whiteflies and their resistant strains, and it is also effective against pests of Coleoptera, Diptera and Lepidoptera, especially for rice The virulence of the stem borer and the three stem borers is higher than that of other neonicotinoid insecticides, and they are widely used in crops such as rice, wheat, vegetables, tobacco, cotton, fruit trees, tea trees, etc., and have the advantages of high efficiency and broad spectrum. In order to prevent potential risks in the application of chlorothialin...

Claims

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Application Information

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IPC IPC(8): C07K7/06C07K7/08G01N33/68
CPCG01N33/68C07K7/06C07K7/08G01N2333/005G01N2430/10G01N2410/00
Inventor 王鸣华华修德丁园
Owner NANJING AGRICULTURAL UNIVERSITY
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