Cationized pleurotus eryngii polysaccharide nanoparticle genetic transmission system and preparation method thereof

A gene transfer system, Pleurotus eryngii polysaccharide technology, applied in the direction of using vectors to introduce foreign genetic material, recombinant DNA technology, etc., can solve the problems of high cost of viral gene vectors, limited gene capacity, carcinogenesis, etc., and achieve good cell adhesion Sex, good binding and release effect, good binding effect

Active Publication Date: 2013-03-13
JIANGSU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although the gene transfection efficiency of the viral gene carrier is relatively high, there are the following problems, such as: can induce host immune response, have potential mutagenic and carcinogenic effects, limited capacity in the gene, preparation of the viral gene carrier The cost is high, especially after the first case of patient death due to the use of adenovirus vectors occurred in 1999, many scientific research institutions in the United States have stopped using viral vectors and focused on the research and application of non-viral vectors

Method used

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  • Cationized pleurotus eryngii polysaccharide nanoparticle genetic transmission system and preparation method thereof
  • Cationized pleurotus eryngii polysaccharide nanoparticle genetic transmission system and preparation method thereof
  • Cationized pleurotus eryngii polysaccharide nanoparticle genetic transmission system and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0033] Example 1. Preparation of Refined Pleurotus eryngii Polysaccharide

[0034] 1. Weigh 1000g of fresh Pleurotus eryngii, crush it into a slurry; extract according to the following process: solid-liquid ratio: 1:4, extraction temperature: 80°C, extraction time 2h / time, extraction times: 2 times; The two extracts were combined, concentrated by rotary evaporation to one-eighth of the volume of the original extract; the concentrated solution was precipitated with 95% absolute ethanol, the final concentration of ethanol was 70%, and stood for 12 hours; the precipitate was collected, and successively washed with absolute ethanol, Wash twice with acetone and ether respectively; vacuum dry to obtain the crude polysaccharide of Pleurotus eryngii.

[0035] 2. Weigh 5g of Pleurotus eryngii crude polysaccharide, dissolve it completely in 50ml double distilled water, add 20% trichloroacetic acid solution to it until the final concentration of trichloroacetic acid is 3%, stir while...

Embodiment 2

[0038] Example 2. Preparation of oxidized Pleurotus eryngii polysaccharide

[0039] Weigh 0.5g refined Pleurotus eryngii polysaccharide, dissolve in 30ml double distilled water, add 0.75g KIO 4 , quickly placed in a dark room, stirred magnetically, and reacted at room temperature for 72 hours; adding 10ml ethylene glycol to the reaction solution to terminate the reaction, and continued the reaction for 30 minutes according to the aforementioned conditions; put the reaction solution into a dialysis bag (cutoff molecular weight > 3500Da), and dialyzed in double distilled water for 48 hours ; The dialysate was freeze-dried to obtain the oxidized Pleurotus eryngii polysaccharide.

Embodiment 3

[0040] Example 3. Preparation of spermine-modified Pleurotus eryngii polysaccharide-DNA plasmid nanocomplex gene delivery system

[0041] A. Take 0.15g oxidized Pleurotus eryngii polysaccharide and dissolve it in 10ml double distilled water; weigh 0.3g spermine and dissolve it in 5ml borate buffer (pH=9); use a disposable syringe to dissolve the borate solution of spermine Slowly added to the Pleurotus eryngii polysaccharide solution, while magnetically stirring; after the addition, magnetically stirred, and reacted at room temperature for 24h; then, added 0.2g sodium borohydride to the reaction solution, and continued to react for 48h under the same conditions; 0.2g sodium borohydride was added to the solution, and the reaction was continued for 24h under the same conditions; the reaction solution was put into a dialysis bag (molecular weight cut-off>3500Da), and dialyzed in double distilled water for 48h; the dialysate was freeze-dried to obtain spermine-modified cationic ...

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Abstract

The invention discloses a cationized pleurotus eryngii polysaccharide and nanoparticle genetic transmission system which is a genetic transmission system of pleurotus eryngii polysaccharide which is modified by amine-group compounds and combined with DNA plasmids. The molecular weight distribution of the pleurotus eryngii polysaccharide is 500-550KD, the mass ratio of the cationized pleurotus eryngii polysaccharide to the DNA plasmids is (0.5-100):1, and the particle size of the cationized pleurotus eryngii polysaccharide-DNA plasmid nanocomposite is 50-100nm. The cationized pleurotus eryngii polysaccharide modified by spermine has the optimal stem cell transfection effect, and the pleurotus eryngii polysaccharide is natural polysaccharide with good biological activity. Based on the natural biological activity of the cationized pleurotus eryngii polysaccharide, the cationized pleurotus eryngii polysaccharide has an additional new biological function, thus being a biodegradable, efficient and harmfulless novel non-viral gene vector material. The invention also discloses the preparation method of the cationized pleurotus eryngii polysaccharide and nanoparticle genetic transmission system.

Description

technical field [0001] The invention relates to the fields of biotechnology and functional polysaccharides of edible fungi, and relates to a gene transfer system, in particular to a cationized Pleurotus eryngii polysaccharide nanoparticle gene transfer system. Background technique [0002] Polysaccharides are macromolecular polymers in which aldose and / or ketose are linked together through glycosidic bonds, which widely exist in nature. They are important components of plants, fungi, and marine organisms. There are a large number of active hydroxyl groups in the molecular structure of polysaccharides, which can carry out many chemical reactions, thereby increasing the biological functions of polysaccharides. [0003] With the development of modern biotechnology and the continuous improvement of the human gene pool, gene therapy has received more and more attention. Various genes related to human diseases have been gradually revealed, making it possible for humans to underst...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/63
Inventor 徐希明邓纹纹余江南王淼曹霞
Owner JIANGSU UNIV
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