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Polymer gene transfection vector based on nucleic acid base derivative and preparation method and application thereof

A nucleic acid base and gene transfection technology, which can be applied to other methods of inserting foreign genetic materials, recombinant DNA technology, etc. It can solve the problems of high cytotoxicity, high material cost, complex synthesis routes, etc. The effect of low cost and simple synthesis process

Inactive Publication Date: 2014-05-21
EAST CHINA NORMAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, gene transfection vectors based on dendrimers have relatively complex synthetic routes, high material costs and high cytotoxicity, which are still the main factors restricting the wide application of these materials in the biomedical field

Method used

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  • Polymer gene transfection vector based on nucleic acid base derivative and preparation method and application thereof
  • Polymer gene transfection vector based on nucleic acid base derivative and preparation method and application thereof
  • Polymer gene transfection vector based on nucleic acid base derivative and preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0046] Preparation and Characterization of Gene Transfection Materials (G5-APu-x, G5-Pu-y, G5-Py-z) Based on Dendrimer and Nucleic Acid Base Derivatives

[0047] (1) if figure 1 The shown gene transfection material G5-APu-x based on dendrimer and nucleic acid base derivatives has a structural formula as shown in the following formula (1A):

[0048]

[0049] Among them, R=G5, is the fifth generation polyamide-amine dendrimer, the central core is ethylenediamine, the terminal group is a primary amine group, n is 6, m is 2; S=APu, is 2- Amino-6-chloropurine; x represents the number of 2-amino-6-chloropurine covalently attached to the surface of the fifth generation polyamido-amine dendrimer;

[0050] Wherein, the structure of G5 is as shown in formula (7A):

[0051]

[0052] In formula (7A), M is the central core of the fifth generation polyamidoamine dendrimer G5: ethylenediamine.

[0053] In a specific embodiment of the present invention, M can also be ammonia, ethylen...

Embodiment 2

[0074] Embodiment 2: Gene transfection experiment of gene transfection material G5-APu-x, G5-Pu-y, G5-Py-z on HEK293 cells

[0075] The gene transfection materials G5-APu-x, G5-Pu-y, G5-Py-z and green fluorescent protein plasmid or luciferase plasmid prepared respectively in Example 1 were formed into a complex at room temperature, and then in HEK293 Transfection was performed on the cells, and the gene transfection efficiency of the material was evaluated by detecting the expression of green fluorescent protein or luciferase. Specific experimental method: Culture HEK293 cells in a 24-well plate and incubate for 24 hours, mix 0.8 micrograms of green fluorescent protein or luciferase plasmid with G5-APu-x, G5-Pu-y, G5-Py-z with different nitrogen and phosphorus After mixing, add to the culture medium and mix evenly; add 500 microliters of culture medium containing 10% serum after incubating with the cells for 6 hours, and treat the cells after 18 hours. The expression of green...

Embodiment 3

[0082] Example 3: Cytotoxicity of gene transfection materials G5-APu-28, G5-Pu-27, G5-Py-16 on HEK293 cells

[0083] To study the cytotoxicity of the gene transfection materials G5-APu-28, G5-Pu-27, and G5-Py-16 in Example 1, the specific experimental method is: incubate HEK293 cells in a 96-well plate, and the cell density is 10 4 Cells / well, remove the medium after culturing for 12 hours, add 100 microliters of fresh medium (containing 10% serum) containing a certain amount of G5-APu-28, G5-Pu-27, and G5-Py-16, the concentration of which is transfection concentration, cultured for 24 hours. Cytotoxicity was detected by MTT assay.

[0084] Experimental results: if Figure 14 The comparison graph of the toxicity of the gene transfection vectors G5-APu-28, G5-Pu-27, G5-Py-16 and other transfection vectors on HEK293 cells shown. Figure 14 It shows that under cell transfection conditions, the gene transfection vectors G5-APu-28, G5-Pu-27, G5-Py-16 have lower cytotoxicity, and...

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Abstract

The invention provides a gene transfection vector based on a dendrimer skeleton and a nucleic acid basic group as shown in a formula (1). The gene transfection vector comprises a dendrimer and a nucleic acid base derivative, wherein the nucleic acid base derivative is in covalent linkage with the surface of the dendrimer. The invention also provides a preparation method of the gene transfection vector shown in the formula (1) and application as a nucleic acid molecule delivery vector. The gene transfection vector provided by the invention is low in synthesis cost and small in cell toxicity, can effectively and safely convey genetic molecules into cells, can be used as the gene transfection vector with the advantages of high efficiency, low toxicity, low cost and the like, and has good application prospect.

Description

technical field [0001] The invention relates to the technical fields of polymer chemistry, organic synthesis and biological materials, in particular to a new polymer gene transfection carrier based on nucleic acid base derivatives and its preparation method and application. Background technique [0002] Gene transfection refers to the process of introducing foreign genes into cells to obtain new genetic traits. Gene transfection vector is the core of gene transfection technology. An ideal gene transfection vector should have the following characteristics: high transfection efficiency, low cytotoxicity, good biological safety, and low price. Currently used gene transfection vectors mainly include viral vectors and non-viral vectors. The main application research still uses viral vectors with high transfection efficiency, but viral vectors have problems such as limited ability to carry genes and potential safety hazards. [0003] Polyamidoamine dendrimers (polyamidoamine, PA...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C08G83/00C08G73/04C12N15/87
Inventor 王辉程义云
Owner EAST CHINA NORMAL UNIV
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