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Application of poloxamer P338 in improving virus infection efficiency of cells and method thereof

A technology of poloxamer, viral infection, applied in the field of viral infection of cells

Pending Publication Date: 2019-06-14
OBIO TECH SHANGHAI CORP LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] However, it has not been disclosed or proven in the art that poloxamers can improve the infection efficiency of viruses (including lentiviruses, adeno-associated viruses) to cells, and at the same time enhance the killing effect on tumor cells

Method used

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  • Application of poloxamer P338 in improving virus infection efficiency of cells and method thereof
  • Application of poloxamer P338 in improving virus infection efficiency of cells and method thereof
  • Application of poloxamer P338 in improving virus infection efficiency of cells and method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0058] Example 1. Poloxamer P338 and lentivirus pLenti-CMV-EGFP-3FLAG co-act on 293T cells

[0059] 1. 293T cell infection

[0060] 1.1 293T cells were plated in 6 wells of a 24-well plate;

[0061] 1.2 Arrange infection when the cell confluency reaches about 75% on the second day; when infecting, according to (number of cells × MOI value / titer of virus stock solution) × 10 3 = virus addition amount (μl), add the corresponding virus volume in each well; add polybrene with a final concentration of 1ug / ml in the polybrene well, and add a final concentration of 1mg / ml in the poloxamer P338 well ml of poloxamer P338; discard the medium after 12-20 hours of infection, and add fresh cell culture medium; take pictures after 48 hours of infection.

[0062] Fluorescence plots showed that poloxamer P338 significantly increased the infection efficiency of lentivirus pLenti-CMV-EGFP-3FLAG on 293T cells at a lower MOI.

[0063] see figure 1 , the infection effect of lentivirus pLenti...

Embodiment 2

[0065] Example 2. Poloxamer P338 and lentivirus pLenti-CMV-EGFP-3FLAG co-act on LLC-MK2 cells

[0066] 1. Infection of LLC-MK2 cells

[0067] 1.1 LLC-MK2 cells were plated in 3 wells of a 24-well plate;

[0068] 1.2 Arrange infection when the cell confluency reaches about 75% on the second day; when infecting, according to (number of cells × MOI value / titer of virus stock solution) × 10 3 = virus addition amount (μl), add the corresponding virus volume to each well; add polybrene with a final concentration of 1 μg / ml to the polybrene well, and add a final concentration of 1 μg / ml to the poloxamer P338 well mg / ml poloxamer P338; discard the culture medium after 12-20 hours of infection, and add fresh cell culture medium; take pictures after 48 hours of infection.

[0069] The fluorescence graph showed that poloxamer P338 significantly improved the infection efficiency of lentivirus pLenti-CMV-EGFP-3FLAG to LLC-MK2 cells at the same MOI.

[0070] join image 3 , the infect...

Embodiment 3

[0071] Example 3: Poloxamer P338 and adeno-associated virus pAAV-CMV-MCS-EGFP-3FLAG co-act on 293T cells

[0072] 1. 293T cell infection

[0073] 1.1 293T cells were plated in 6 wells of a 24-well plate;

[0074] 1.2 Arrange infection when the cell confluency reaches about 75% on the second day; when infecting, according to (number of cells × MOI value / titer of virus stock solution) × 10 3 = virus addition (μl), add the corresponding virus volume to each well; add poloxamer P338 with a final concentration of 1mg / ml to the poloxamer P338 well; discard the medium after 12-20h of infection, Fresh cell culture medium was added; photographs were taken 48h after infection.

[0075] The fluorescence graph showed that poloxamer P338 significantly improved the infection efficiency of adeno-associated virus PAAV-CMV-MCS-EGFP-3FLAG on 293T cells.

[0076] join Figure 4 , Adeno-associated virus pAAV-CMV-MCS-EGFP-3FLAG at MOI=1×10 3 Infection effect diagram of 293T cells under the ...

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Abstract

The invention discloses a method for utilizing poloxamer P338 to improve the virus infection efficiency of cells and application thereof. The use of the poloxamer P338 can increase the infection efficiency of lentivirus at low MOI to 293T cells, LLC-MK2 cells and the like. The use of the poloxamer P338 can also promote the infection of adeno-associated virus at the same MOI value to the 293T cellsto enhance the expression of foreign genes carried by viral vectors.

Description

technical field [0001] The invention belongs to the technical field of virus-infected cells, and more specifically, the invention relates to an application and a method for improving the efficiency of virus-infected cells based on the use of poloxamer P338. Background technique [0002] The use of exogenous nucleic acid substances carried by lentiviral vectors and adeno-associated virus vectors to edit eukaryotic cell genomes has become an effective means for studying gene functions and precise gene therapy, and has been widely used in basic research and some clinical research stages. However, the infection efficiency of viruses to cells, especially primary cells, and to cells in individuals is still low, and there are varying degrees of cell dependence. In addition, the use of high-dose virus to improve infection efficiency not only increases the complexity and usage of virus packaging, but also increases the actual cost of use, and the invasion of a large number of foreign...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/867C12N15/864
Inventor 刘晓芬杨兴林杨蕊菊夏清梅鲁济真
Owner OBIO TECH SHANGHAI CORP LTD
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