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Phage display polypeptide specifically binding to imidacloprid antibody and application thereof

An imidacloprid, specific technology, applied in the fields of peptides, material testing products, biological tests, etc., can solve the problems of unseen peptides, etc., and achieve the effects of high accuracy, high sensitivity and strong specificity

Inactive Publication Date: 2019-09-20
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 studies and reports on polypeptides that specifically bind to imidacloprid antibodies and their applications.

Method used

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  • Phage display polypeptide specifically binding to imidacloprid antibody and application thereof
  • Phage display polypeptide specifically binding to imidacloprid antibody and application thereof
  • Phage display polypeptide specifically binding to imidacloprid antibody and application thereof

Examples

Experimental program
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Embodiment 1

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

[0029] 1. Panning of phage clones that specifically bind to imidacloprid 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 imidacloprid antibody purified by a 100 μg / mL protein A column 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) 3 times with PBST, add 300 μL of 5% skimmed milk powder to each well, incubate at 37°C for 2 hours, wash 3 times with PBST, and store at 4°C for later use;

[0034] (4) Add 100 μL 2×10 11 Phage (containing 5% skimmed milk powder), slightly shaken at room temperature for 1 hour;

[0035] (5) Take 20mL of LB med...

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Abstract

The invention belongs to the biotechnology field, and relates to a polypeptide specifically binding to an imidacloprid antibody and an application of the polypeptide in detection of imidacloprid. A phage display technology is adopted, a phage display random linear dodecapeptide library is screened three times by the imidacloprid antibody purified by a protein A column, and phage clones that bind to the imidaclone antibody are screened out; a number of phage clones are randomly picked up for amplification and plasmid extraction, positive phage clones are selected by a phage ELISA method, and the positive clones are sequenced to obtain a polypeptide sequence. The invention also relates to the application of the phage display polypeptide in detection of imidacloprid. The phage enzyme-linked immunosorbent assay method established by the phage display polypeptide can be used for rapid, sensitive, simple and cheap detection of imidacloprid residues in environment and agricultural products.

Description

technical field [0001] The invention belongs to the field of biotechnology, and relates to a polypeptide with specific binding to imidacloprid antibody, including the application of the polypeptide in detecting imidacloprid. Background technique [0002] Imidacloprid is a super-efficient neonicotinoid insecticide with multiple effects such as contact killing, stomach toxicity and systemic absorption. Due to the extensive, continuous and large-scale use of imidacloprid, the death of many natural enemies of pests has been caused. Studies have found that imidacloprid is highly toxic to the natural enemies of rice planthoppers, Lygus spp. Bees, bees, and beneficial organisms in the soil can all cause damage. In order to prevent the potential risks of imidacloprid application, a sensitive, rapid and selective residue detection method is needed. [0003] At present, for the detection of imidacloprid residues, instrumental analysis methods (such as liquid chromatography, gas chro...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K7/08G01N33/68
CPCC07K7/08G01N33/68G01N33/94
Inventor 王鸣华华修德杜梅丁园陈贺
Owner NANJING AGRICULTURAL UNIVERSITY
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