Modified polyethyleneimin, preparation method of modified polyethyleneimine, gene transfection reagent and application of gene transfection reagent

A technology of polyethylenimine and gene transfection, applied to other methods of inserting foreign genetic material, using vectors to introduce foreign genetic material, recombinant DNA technology, etc., can solve unfavorable production scale and commercial promotion, low cytotoxicity, and product poor repeatability

Active Publication Date: 2015-03-18
SHENZHEN INST OF ADVANCED TECH CHINESE ACAD OF SCI
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  • Abstract
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  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, in the existing methods for chemically modifying PEI, some steps are relatively cumbersome, which is not conducive to expanding production scale and commercial promotion, and the cytotoxicity of the obtained modified PEI has decreased, but the transfection performance has not been improved. The order of magnitude increases; some require high-temperature reactions greater than 100°C, and strict anhydrous and oxygen-free harsh conditions are required during the reaction, and some of the ring-opening polymerization modification methods have a molecular weight distribution due to the reaction mechanism of the ring-opening polymerization itself, which makes it impossible to achieve molecular weight. Precise regulation, so it is easy to cause poor product repeatability and it is difficult to ensure high transfection efficiency and low cytotoxicity

Method used

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  • Modified polyethyleneimin, preparation method of modified polyethyleneimine, gene transfection reagent and application of gene transfection reagent
  • Modified polyethyleneimin, preparation method of modified polyethyleneimine, gene transfection reagent and application of gene transfection reagent
  • Modified polyethyleneimin, preparation method of modified polyethyleneimine, gene transfection reagent and application of gene transfection reagent

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preparation example Construction

[0051] The preparation method of the modified polyethylenimine of one embodiment, comprises the steps:

[0052] Add polyethyleneimine and ε-caprolactone to the solvent at a mass ratio of 1:0.01 to 2.7, and add a catalyst to obtain a mixture, stir the mixture at 15°C to 35°C for 5 minutes to 30 minutes, and then heat to Boiling, stirring for 24 hours to 72 hours, separation and purification to obtain modified polyethyleneimine.

[0053] The polyethyleneimine is a branched polyethyleneimine or a linear polyethyleneimine.

[0054] The solvent is a solvent that can dissolve polyethyleneimine and ε-caprolactone, preferably anhydrous chloroform (CHCl 3 ). Preferably, the mass ratio of polyethyleneimine to solvent is 1:10-200.

[0055] Catalyst is the catalyst that can catalyze this reaction, is preferably stannous octanoate (Sn(Oct) 2 ). The mass ratio of polyethyleneimine to the catalyst is preferably 1:0.0005-0.01.

[0056] Stir the mixture of polyethyleneimine, ε-caprolacto...

Embodiment 1

[0070] 1. Preparation of modified polyethyleneimine

[0071] First, dissolve 1 part by mass of branched PEI (expressed as bPEI25K) with a weight average molecular weight of 25000 g / mol in 10 parts by mass of anhydrous chloroform, then add 0.01 part by mass of ε-caprolactone at room temperature, and then add 0.0005 parts by mass of stannous isooctanoate to obtain a mixture, the mixture was stirred at 23°C for 30 minutes, then heated to 76°C, the mixture was boiled, stirred and reacted for 48 hours, then filtered, and the residue was taken to obtain an insoluble chloroform solution The crude product of branched polyethyleneimine modified by ε-caprolactone, the crude product of branched polyethyleneimine modified by ε-caprolactone is dissolved in 5 parts by mass of methanol solution, dropwise at room temperature and under stirring Add 50 parts by mass of anhydrous ether, filter, and the obtained precipitate is modified polyethyleneimine, expressed as bPEI25K-CL.

[0072] The str...

Embodiment 2

[0097] Preparation of Modified Polyethyleneimine

[0098] Dissolve 1 part by weight of branched PEI (bPEI75K) with a weight average molecular weight of 75,000 g / mol in 200 parts by weight of anhydrous chloroform, then add 2.7 parts by weight of ε-caprolactone at room temperature, and then add 0.01 parts by weight Stannous isooctanoate to obtain a mixture, the mixture was stirred at 35°C for 5 minutes, then heated to 76°C, the mixture was boiled, stirred and reacted for 72 hours, then filtered, and the residue was taken to obtain ε insoluble in chloroform solution -The crude product of branched polyethyleneimine modified by caprolactone, the crude product of branched polyethyleneimine modified by caprolactone is dissolved in 10 parts by weight of methanol solution, and 100 parts by weight is added dropwise at room temperature and under stirring conditions Anhydrous ether, filtered, the resulting precipitate is modified polyethyleneimine.

[0099] The structural formula of this...

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Abstract

The invention relates to modified polyethyleneimine, a preparation method of modified polyethyleneimine, a gene transfection reagent and application of the gene transfection reagent. The modified polyethyleneimine comprises polyethyleneimine as a main chain and epsilon-caprolactone grafted onto polyethyleneimine, epsilon-caprolactone and primary amine amino group or secondary amine amino group of polyethyleneimine are subjected to coordination ring-opening grafting reaction and the epsilon-caprolactone is not subjected to coordination ring-opening polymerization reaction. The experiment shows that modified polyethyleneimine with the structure has relatively high gene transfection efficiency which is far higher than those of commercially available products branched polyethylenimine and liposomes 2000, and compared with branched polyethyleneimine having weight-average molecular weight of 25000g / mol, no significant cytotoxicity is observed in modified polyethyleneimine. Therefore, modified polyethyleneimine can be considered as an excellent gene delivery system.

Description

technical field [0001] The invention relates to the technical field of gene transfection biology, in particular to a modified polyethylenimine and a preparation method thereof, a gene transfection reagent and an application thereof. Background technique [0002] Gene therapy (gene therapy) and gene modification (gene modification) refer to the introduction of exogenous functional genes into target cells to correct, compensate or modify the original gene sequence of the organism, and change the gene expression function to meet experimental needs and disease treatment the goal of. However, exogenous genes cannot effectively enter target cells by themselves, and gene delivery systems are needed to achieve gene expression. As a tool for carrying genes, the gene delivery system can deliver the target gene into the target cells, so as to exert the specific function of the target gene. [0003] Currently, there are two broad categories of gene delivery systems: viral and nonviral...

Claims

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Application Information

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IPC IPC(8): C08G73/04C12N15/87C12N15/85C12N5/07
Inventor 王志勇谢丽斯刘新郑海荣
Owner SHENZHEN INST OF ADVANCED TECH CHINESE ACAD OF SCI
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