Recombinant expression method of human cryptochrome protein I (hcry1) and its application in preparation of radiotherapy protective agent

A cryptochrome and protective agent technology, which is applied to the recombinant expression of human cryptochrome protein I (hCRY1) and its application in the preparation of radiotherapy protective agents, can solve the problems of lack of application prospects, high cost of peptide synthesis, and lack of literature Report on animal CRY1 protein prokaryotic expression and purification methods and other issues

Active Publication Date: 2017-03-22
黄文林 +1
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Problems solved by technology

However, the cost of peptide synthesis is too high, and it lacks application prospects as a drug production
So far, no literature has reported the prokaryotic expression and purification method of animal CRY1 protein

Method used

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  • Recombinant expression method of human cryptochrome protein I (hcry1) and its application in preparation of radiotherapy protective agent
  • Recombinant expression method of human cryptochrome protein I (hcry1) and its application in preparation of radiotherapy protective agent
  • Recombinant expression method of human cryptochrome protein I (hcry1) and its application in preparation of radiotherapy protective agent

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Embodiment

[0029] The following description of the present invention is based on the examples to further illustrate the present invention, the present invention is not limited to the following examples. For the embodiment of the method of the present invention as follows:

[0030] 1. Construction of prokaryotic expression plasmid pET28a(+)-hCRY1:

[0031] Using the hCRY1 eukaryotic expression plasmid pcDNA3.1-hCRY1 stored in the laboratory as a template, design specific primers according to the sequence of the hCRY1 coding region: the upstream primer is hCRY1FW (5'-ATACTCGAGGCTAAGCCTTCC-3'), and the downstream primer is hCRY1RV (5' -CGCGGATCCTACGTTATACT-3') (both upstream and downstream primers were synthesized by Beijing Aoke Dingsheng Biotechnology Co., Ltd.). PCR amplifies the required target fragment (total system 25uL, 94°C pre-denaturation for 3min, 32 cycles: 94°C for 30s, 55°C for 30s, 72°C for 2min, and finally 72°C for 10 minutes). Use restriction endonucleases XhoI and BamlI...

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Abstract

The invention relates to a recombinant expression method for a human cryptochrome protein I (hCRY1) and an application of the hCRY1 in the preparation of a radiotherapy protective agent. The method comprises the steps of recombinant expression of the protein hCRY1 in a prokaryotic organism, purification, activation, functional detection and the like. According to the method, about 20 to 25 milligrams of hCRY1 can be averagely obtained from every liter of bacterial liquid, the yield is far higher than that of any other conventional method, and the cost is calculated to be far lower than that of a eukaryotic expression and purification method and a polypeptide chemical synthesis method. Moreover, a protein product obtained by the method is proved to have ray absorption and radiation injury protection functions, is an active protein potential to be developed into a radiotherapy protection medicament, and can be used as the radiotherapy protective agent.

Description

technical field [0001] The invention relates to a recombinant expression method of hCRY1 protein and its application in preparing radiotherapy protective agent. Background technique [0002] Cancer is the greatest enemy of human health, and the number of people who die of cancer in the world is close to 10 million every year. In the clinical treatment of cancer, surgery, radiotherapy and chemotherapy are the three most important treatment methods. Among them, radiotherapy is more selective because of its broad indications. Radiation therapy is required. [0003] The purpose of radiotherapy is to concentrate the radiation dose to the lesion area (target area) to the maximum extent, and kill tumor cells, while the surrounding normal tissues or organs receive less or no unnecessary irradiation. Some important organs such as brainstem, lens , spinal cord, kidneys, gonads, etc., need special protection. Neither traditional radiotherapy nor modern precision radiotherapy can ful...

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/70C07K14/47A61K38/17A61P17/16
Inventor 潘长穿黄文林姚辰陈帅
Owner 黄文林
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