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Transient transfection reagent and use thereof

A technology of transient transfection and transfection reagent, which is applied in the field of gene transfection and achieves the effects of convenient and quick transfection, high solution stability and low cost.

Inactive Publication Date: 2017-04-26
JILIN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Aiming at the deficiencies of existing transfection reagents, the present invention provides a transient transfection reagent and its application method, which have the advantages of high and stable transfection efficiency, low cytotoxicity, simple and time-saving operation, stable solution, and a wide range of adaptability to host cells, etc.

Method used

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  • Transient transfection reagent and use thereof
  • Transient transfection reagent and use thereof
  • Transient transfection reagent and use thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0020] Example 1 Transfection

[0021] (1) First add 150 μL of OptiMEM medium to the centrifuge tube, then add 3 μg of GFP plasmid, and then follow the plasmid (μg): transfection reagent (μL) 1:1.5, 1:2, 1:3, 1:4, 1: 6 Add PEI-max transfection reagent. The commercial transfection reagent lipofectamine 3000 was used as a control for reference.

[0022] (2) Incubate at room temperature for 5 minutes and add to DMEM cell culture medium;

[0023] (3) Test after 24-36 hours.

[0024] Experimental results such as figure 1 As shown, it can be seen that the ratio of plasmid to transfection reagent is 1:3: when the transfection effect is the best. The transfection effect was close to the commercial transfection reagent lipofectamine 3000.

Embodiment 2

[0025] Example 2 Co-transfection

[0026] Co-transfection plasmids: 1.5 μg each of GFP and mcherry plasmids; the specific transfection scheme is the same as in Example 1.

[0027] Experimental results such as figure 2 As shown, it can be seen from the comparison chart of GFP and mCherry co-transfection that both proteins are expressed in large quantities under the three transfection conditions.

Embodiment 3

[0028] Example 3 Comparison of protein co-expression with commercial transfection reagents:

[0029] 6 μg of DNA (3 μg each of GFP plasmid and mcherry plasmid) was transfected in HEK293T cells, and the specific transfection method was the same as in Example 1.

[0030] from image 3 It can be seen from the results that the co-transfection efficiency of PEI-max reagent combined with the optimized transfection protocol can achieve similar effects to commercial transfection reagents.

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Abstract

The invention provides a transient transfection reagent and a use thereof. The transfection efficiency of the transfection reagent is high and stable and is equivalent to the transfection efficiency of Lipofectamine 3000 (hereinafter referred to as lipo3000). The transient transfection reagent has high transfection efficiency both in plasmid single transfection and co-transfection and has very small cytotoxicity. The transient transfection reagent can be simply operated. Compared with lipo3000, the transient transfection reagent is free of plasmid dilution and reagent transfection, realizes transfection in 10min, has high solution stability, can be stored at a temperature of 4 DEG C for a long time, has a low cost, very high transfection efficiency and high stability, can be operated easily, is suitable for a wide range of host cells, is not influenced by antibiotics of the culture medium in transfection efficiency and is an excellent choice for a lab where a lot of transient transfection experiments are carried out.

Description

technical field [0001] The invention relates to the technical field of gene transfection, in particular to a transient transfection reagent and an application method thereof. Background technique [0002] Transfection requires certain transfection reagents to transport the vector carrying the gene of interest into the host cell. At present, the most commonly used transfection reagents are cationic liposomes and cationic polymers, which have characteristics similar to viruses and can easily penetrate cell membranes. Among them, cationic liposomes have high efficiency in gene transfection in vitro, however, in vivo, it is rapidly cleared by serum, accumulates in lung tissue, and induces a strong anti-inflammatory response, which will lead to high levels of toxicity, so , which limits its application to a large extent. Due to the limitations of cationic liposomes, cationic polymeric transfection reagents are gaining increasing attention. [0003] This reagent uses a cationic...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/63C12N15/85
CPCC12N15/63C12N15/85C12N2800/107
Inventor 杨亮左泽乘莫伟亮姚楠朴明鑫边鸣镝杨振明
Owner JILIN UNIV
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