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Bombinin-8, gene of Bombinin-8 and application of Bombinin-8 in pharmaceuticals

A technology of biologically active peptides and genes, applied in the field of biomedicine, can solve problems such as the lack of reports on functional research, and achieve the effects of convenient chemical synthesis and genetic engineering preparation, significant anti-tumor activity, and simple structure

Inactive Publication Date: 2017-05-31
TIANJIN MEDICAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Due to the limitation of the geographical distribution of Bombina orientalis, there are not many reports on the active substances in the secretion of Bombina orientalis and their functional research in the world.

Method used

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  • Bombinin-8, gene of Bombinin-8 and application of Bombinin-8 in pharmaceuticals
  • Bombinin-8, gene of Bombinin-8 and application of Bombinin-8 in pharmaceuticals
  • Bombinin-8, gene of Bombinin-8 and application of Bombinin-8 in pharmaceuticals

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0023] Extraction of Bombina orientalis skin secretions:

[0024] Select adult oriental bombinas (4-6cm long, half male and half male, n=10), rinse them with ultrapure water, and then gently massage the skin surface of the back of the bombina. Gentle massage was applied for several minutes, and a large amount of white viscous foamy exudate was secreted from the skin glands on the back. The skin secretion was washed with ultra-pure water into a clean container, and it was quickly frozen in liquid nitrogen and placed in a freeze dryer. Freeze-dried and stored at -20°C.

Embodiment 2

[0026] Molecular Cloning of Bioactive Peptide Bombinin-8 from Bombina orientalis

[0027] 1. Extraction of RNA from Bombina orientalis secretion

[0028] When amphibians secrete defensive secretions, the skin at the glands on the back will be slightly damaged, and the RNA in the skin cells will be released into the secretions at the same time. Many experiments have proved that the RNA of amphibians can be detected in the secretions. Therefore, we used the method of isolating RNA from secretions to extract total RNA from Bombina orientalis skin. The specific steps are as follows: (1) The RNA in Bombina orientalis secretions was extracted using MiniBEST Universal RNA from TaKaRa Company

[0029] Extraction Kit.

[0030] (2) First weigh 5 mg of freeze-dried Bombina orientalis skin secretion, add it to a 1.5 mL sterilized tube containing 500 μL of lysis solution Buffer RL, and repeatedly pipette until there is no obvious precipitation in the lysis solution. Centrifuge the lysate...

Embodiment 3

[0058] Separation, purification and amino acid sequence determination of biologically active peptides from Bombina orientalis:

[0059] 1. Separation and purification of biologically active peptides from Bombina orientalis

[0060] The freeze-dried skin secretion of Bombina orientalis in Example 1 was dissolved in a certain volume of Buffer A (TFA / water (0.05 / 99.95, V / V)), and centrifuged at 3000 g for 10 minutes to remove insoluble matter. The supernatant was separated and purified by reverse-phase semi-preparative HPLC with a C5 chromatographic column, eluted with a gradient of 0-70% acetonitrile, and the sample eluate was collected by the Frac-920 automatic fraction collector with an interval of 1 minute. The result is as figure 1 As shown, the arrow points to the biologically active peptide Bombinin-8 of Bombina orientalis.

[0061] 2. Structural Determination of Bioactive Peptides from Bombina orientalis

[0062] The molecular weight of the polypeptide was determined b...

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Abstract

The invention relates to Bombinin-8, a gene of the Bombinin-8 and application of the Bombinin-8 in pharmaceuticals. The Bombinin-8 provided by the invention has broad-spectrum antibacterial activity, plays a role in inhibiting the growth and propagation of three kinds of liver cancer cells and can be applied to the preparation of antitumor pathogenic microorganism infection resisting drugs.

Description

technical field [0001] The invention relates to a bioactive peptide Bombinin-8 of Bombina orientalis and its gene and application in pharmacy, belonging to the field of biomedicine. Background technique [0002] Malignant tumors are a class of diseases that seriously threaten human health and life. Although traditional chemotherapy can inhibit the development of malignant tumors, the antitumor drugs used in the chemotherapy process not only kill tumor cells, but also have relatively large lethal and toxic side effects on normal cells. In clinical use, tumor drug resistance may also occur. Therefore, the study of new anti-tumor drugs with high efficiency and low toxicity has become an important problem to be solved urgently in today's tumor treatment. Liver cancer is one of the top ten malignant tumors in the world, and the incidence rate of liver cancer in my country is the second among malignant tumors. The recurrence rate of liver cancer is high, and traditional chemoth...

Claims

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Application Information

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IPC IPC(8): C07K14/46C12N15/12A61K38/17A61P31/04A61P31/10A61P35/00
CPCA61K38/00C07K14/463
Inventor 王冉孔德新周畅彭鑫
Owner TIANJIN MEDICAL UNIV
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