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Humanized butyrylcholine esterase (H-Bche), coding gene thereof and applications of H-Bche and coding gene

A human-sourced butyrylcholine and butyrylcholine-sourced technology, applied in applications, genetic engineering, plant genetic improvement, etc., can solve the problems of low yield and activity, long cycle time and high cost of butyrylcholinesterase

Active Publication Date: 2014-10-22
BEIJING UNIV OF AGRI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] The purpose of the present invention is to overcome the defects of long cycle, low yield and activity, and high cost of the existing preparation of butyrylcholinesterase, and to provide a human-sourced butyrylcholinesterase with high activity, high yield, and low cost. Alkaline esterase method

Method used

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  • Humanized butyrylcholine esterase (H-Bche), coding gene thereof and applications of H-Bche and coding gene
  • Humanized butyrylcholine esterase (H-Bche), coding gene thereof and applications of H-Bche and coding gene
  • Humanized butyrylcholine esterase (H-Bche), coding gene thereof and applications of H-Bche and coding gene

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Experimental program
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Effect test

Embodiment 1

[0042] 1. Obtaining the sequence shown in SEQ ID NO.1

[0043] 1.1 Acquisition and amplification of the nucleotide sequence shown in SEQ ID NO.2

[0044] The artificially synthesized 5' end has restriction site GAATTC and His tag (CATCATCATCACCATCAT), and the 3' end has the nucleotide sequence shown in SEQ ID NO.2 of restriction site GCGGCCGC. The sequence shown in SEQ ID NO.2 was connected to the PUC57 vector, and transformed into Escherichia coli competent cell DH5α.

[0045] The Escherichia coli introduced with the PUC57 vector linked with the sequence shown in SEQ ID NO.3 was cultured in Amp-resistant LB liquid medium at 150 rpm and pH 7.0 for 12 hours.

[0046] The plasmid was extracted with the above cultured bacterial solution, extracted according to the instructions of the plasmid mini-extraction kit (purchased from Beijing Tiangen Biotechnology Co., Ltd., product number: DP103), and the plasmid was verified by 1% agarose electrophoresis. The verification result showe...

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Abstract

The invention discloses a coding gene of butyrylcholine esterase (Bche). The gene has a nucleotide sequence shown in SEQ ID No:1 or a nucleotide sequence which can still carry out coding after carrying out substitution, deletion or addition of one or a plurality of nucleotides on the nucleotide sequence shown in SEQ ID No:1 and has the functions of Bche. The invention also provides a recombinant vector and transgenic cell containing the coding gene and applications of the recombinant vector and the transgenic cell. The coding gene has the beneficial effects that Bche can be quickly produced by preferably expressing the gene for coding Bche in a pichia pastoris cell; besides, Bche can be efficiently produced in quantity only by culturing the cell containing the Bche gene; seen from the embodiment, the highest activity of Bche prepared by the method can be 0.6789pmol / mg.min.

Description

technical field [0001] The present invention relates to a gene encoding human butyrylcholinesterase, a recombinant vector and a transgenic cell containing the encoding gene, and human butyryl cholinesterase encoded by the gene encoding human butyrylcholinesterase Cholinesterase, in addition, the present invention also relates to the application of the encoding gene, the recombinant vector, the transgenic cell and the human butyrylcholinesterase in the detection of organophosphorus and carbamate pesticides. Background technique [0002] The safety of food and agricultural products is a major global issue related to the national economy and people's livelihood, among which the problem of excessive pesticide residues in agricultural products is a hot issue of social concern. The high toxicity and high risk of organophosphorus and carbamate pesticides have been focused on by the state and relevant departments. my country has successively banned HCH, DDT, toxaphene, dibromochlor...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/55C12N15/81C12N1/19C12N9/18C12Q1/46C12R1/84
Inventor 陈湘宁谢远红岳溪许丽田晶晶
Owner BEIJING UNIV OF AGRI