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Cationic transfection agent and preparation and application methods thereof

A transfection reagent, a cationic technology, applied in the field of genetic engineering, can solve problems such as inability to pass on time, time-consuming transfection operation, and increase in cell survival rate, so as to reduce cytotoxicity, optimize components, and improve Effect of transfection efficiency

Active Publication Date: 2019-08-16
苏州博特龙免疫技术有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although transfection reagents are constantly being improved, there are still certain defects, such as: low transfection efficiency; cell survival rate after transfection still needs to be improved; Transfect immediately when the parietal cells have not adhered to the wall after passage); the transfection operation takes a long time, etc.

Method used

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  • Cationic transfection agent and preparation and application methods thereof
  • Cationic transfection agent and preparation and application methods thereof
  • Cationic transfection agent and preparation and application methods thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] Embodiment 1: A cationic transfection reagent and its preparation

[0042] A kind of preparation method of cationic transfection reagent comprises the following steps:

[0043] (1) Dissolve polyethyleneimine and cationic polyacrylamide in PBS buffer in proportion, stir at room temperature for 15 minutes, mix evenly, and filter through a 0.2 μm filter membrane to obtain mixed solution 1, in which polyethyleneimine and cationic polyacrylamide The concentrations of acrylamide were 10ng / mL and 20ng / mL, respectively;

[0044] (2) Didodecyldimethylammonium bromide and vitamin D were dissolved in 85% ethanol aqueous solution in proportion, stirred at room temperature for 20 minutes, mixed evenly, and filtered with a 0.2 μm filter membrane to obtain a mixed solution 2, wherein The concentrations of didodecyldimethylammonium bromide and vitamin D were 20ng / mL and 10ng / mL, respectively;

[0045] (3) Mix the mixed solution 1 obtained in step (1) and the mixed solution 2 obtained...

Embodiment 2

[0047] Embodiment 2: A cationic transfection reagent and its preparation

[0048] A kind of preparation method of cationic transfection reagent comprises the following steps:

[0049] (1) Dissolve polyethyleneimine and cationic polyacrylamide in PBS buffer in proportion, stir at room temperature for 15 minutes, mix evenly, and filter through a 0.2 μm filter membrane to obtain mixed solution 1, in which polyethyleneimine and cationic polyacrylamide The concentrations of acrylamide were 20ng / mL and 10ng / mL respectively;

[0050] (2) Dissolve dioleoylphosphatidylethanolamine and vitamin D in 85% ethanol aqueous solution in proportion, stir at room temperature for 20 minutes, mix well, and filter with a 0.2 μm filter membrane to obtain mixed solution 2, in which dioleoylphosphatidylethanolamine The concentrations of ethanolamine and vitamin D were 40 ng / mL and 4 ng / mL, respectively;

[0051] (3) Mix the mixed solution 1 obtained in step (1) and the mixed solution 2 obtained in s...

Embodiment 3

[0053] Embodiment 3: A kind of cationic transfection reagent and its preparation

[0054] A kind of preparation method of cationic transfection reagent comprises the following steps:

[0055] (1) Dissolve polyethyleneimine and cationic polyacrylamide in PBS buffer in proportion, stir at room temperature for 15 minutes, mix evenly, and filter through a 0.2 μm filter membrane to obtain mixed solution 1, in which polyethyleneimine and cationic polyacrylamide The concentrations of acrylamide were 30ng / mL and 6ng / mL, respectively;

[0056] (2) Dioctadecylaminoglycylcarboxyspermine and vitamin D were dissolved in 85% ethanol aqueous solution in proportion, stirred at room temperature for 20 minutes, mixed evenly, and filtered with a 0.2 μm filter membrane to obtain the mixed solution 2, Wherein the concentrations of dioctadecylaminoglycyl carboxyspermine and vitamin D are 60ng / mL and 2ng / mL respectively;

[0057] (3) Mix the mixed solution 1 obtained in step (1) and the mixed solu...

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Abstract

The invention provides a cationic transfection agent and preparation and application methods thereof and relates to the field of genetic engineering. The cationic transfection agent is composed of polyethyleneimine, cationic polyacrylamide, cationic liposome, fatty alcohol polyoxyethylene ether ammonium sulfate and vitamin D. By optimizing the components, the cationic transfection agent can significantly improve the transfection efficiency and achieve a 293FT cell transfection rate up to 95%. Besides, the cationic transfection agent can achieve transfection instantly when adherent cells are not adhered after passage, thereby achieving instant passage and transfection; transfection operation is simple and rapid and can be applied within one minute. Therefore, the cationic transfection agentcan be widely applied to rapid, large-scale and instant transfection of various mammalian cells.

Description

technical field [0001] The invention relates to the field of genetic engineering, in particular to a cationic transfection reagent and its preparation method and application. Background technique [0002] Cell transfection is a technique for introducing exogenous genes into eukaryotic cells. It is widely used in protein expression localization, gene function research, gene expression regulation, and gene therapy. It is an important tool in the field of genetic engineering. The methods of cell transfection can be divided into three categories: physical, chemical and biological. Among them, the physical mediation methods include electroporation, microinjection and gene gun, etc.; the chemical mediation methods include cationic liposome method, calcium phosphate co-precipitation method and cationic polymer method; the biological mediation method is mainly virus mediation transfection method. Although the virus-mediated transfection method has the advantages of high transfecti...

Claims

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Application Information

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IPC IPC(8): C12N15/85
CPCC12N15/85
Inventor 刘立成
Owner 苏州博特龙免疫技术有限公司
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