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Culture medium additive for promoting protein expression and application thereof

A protein expression and culture medium technology, applied in the culture process, tissue culture, microorganisms, etc., can solve the problems of virus infection, complex composition, low expression yield, etc., and achieve the effect of increasing yield

Pending Publication Date: 2021-07-23
内蒙古金源康生物工程股份有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The exogenous proteins produced by mammalian cells after translation and modification are far superior to prokaryotic expression systems and eukaryotic expression systems such as yeast and insect cells in terms of activity, and are closer to natural proteins, but they are complex in composition and require high operating techniques. The expression yield is not large, the yield is low, and sometimes it will cause virus infection, etc., which are the shortcomings of this expression system

Method used

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  • Culture medium additive for promoting protein expression and application thereof
  • Culture medium additive for promoting protein expression and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0034] Control medium: commercial medium 302 culture (Millipore Sigma, 24571C).

[0035] Test group culture medium: add supplements on DMEM / F12 basic medium, supplement components and working concentration: vitamin C10μM, ethanolamine 10μM, L-asparagine 10mg / L, L-cysteine ​​hydrochloride 5mg / L , L-cystine dihydrochloride 10mg / L, L-proline 20mg / L, L-tryptophan 10mg / L, soybean peptone 1g / L, N-acetylcysteine ​​2mM, pluronic F681g / L.

[0036] Suspended BHK-21 cells were cultured in the culture medium of the control group and the medium of the experimental group, respectively, in Corning 125 shake flasks, 37°C, 8% CO 2 , cultured on a shaker at 130rpm, when the cell density reached 2×10 6 cells / ml, it can be subcultured, and the subculture ratio is 1:4. When the cell density reaches 2×10 6 cells / ml for cell plasmid transfection, the concentration of the constructed fusion expression vector was transfected at 1.5×10 per 18 μg of plasmid DNA. 7 BHK-21 cells diluted to 30ml of ...

Embodiment 2

[0038] Control medium: CHO cells, BHK-21 cells and 293 cells in commercial medium 302 culture (Millipore Sigma, 24571C).

[0039] Test group culture medium: add supplements on DMEM / F12 basic medium, supplement components and working concentration: vitamin C 50μM, ethanolamine 50μM, L-asparagine 50mg / L, L-cysteine ​​hydrochloride 50mg / L L, L-cystine dihydrochloride 50mg / L, L-proline 60mg / L, L-tryptophan 50mg / L, soybean peptone 5g / L, N-acetylcysteine ​​10mM, Pram Nick F685g / L.

[0040] Suspended BHK-21 cells were cultured in the culture medium of the control group and the medium of the experimental group, respectively, in Corning 125 shake flasks, 37°C, 8% CO 2 , cultured on a shaker at 130rpm, when the cell density reached 2×10 6 cells / ml, it can be subcultured, and the subculture ratio is 1:4. When the cell density reaches 2×10 6 cells / ml for cell plasmid transfection, the concentration of the constructed fusion expression vector was transfected at 1.5×10 per 18 μg of ...

Embodiment 3

[0042] Control medium: commercial medium 302 culture (Millipore Sigma, 24571C).

[0043] Experimental group culture medium: add supplements on DMEM / F12 basic medium, supplement components and working concentration: vitamin C 20μM, ethanolamine 25μM, L-asparagine 30mg / L, L-cysteine ​​hydrochloride 30mg / L L, L-cystine dihydrochloride 35mg / L, L-proline 35mg / L, L-tryptophan 25mg / L, soybean peptone 2g / L, N-acetylcysteine ​​5mM, Pram Nick F68 1.5g / L.

[0044] Suspended BHK-21 cells were cultured in the culture medium of the control group and the medium of the experimental group, respectively, in Corning 125 shake flasks, 37°C, 8% CO 2 , cultured on a shaker at 130rpm, when the cell density reached 2×10 6 cells / ml, it can be subcultured, and the subculture ratio is 1:4. When the cell density reaches 2×10 6 cells / ml for cell plasmid transfection, the concentration of the constructed fusion expression vector was transfected at 1.5×10 per 18 μg of plasmid DNA. 7 BHK-21 cells in 30m...

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Abstract

The invention discloses a culture medium additive for promoting protein expression, and belongs to the technical field of cell culture. The culture medium additive comprises vitamin C, ethanolamine, L-asparagine, L-cysteine hydrochloride, L-cystine dihydrochloride, L-proline, L-tryptophan, soy peptone, N-acetylcysteine and pluronic F68. The culture medium additive for promoting protein expression is added into DMEM, DMEM / F12 and RPMI1640 basic culture media, so that the yield of cell expression protein can be increased, and a foundation is laid for improving protein antibody drugs.

Description

technical field [0001] The invention belongs to the technical field of cell culture, and in particular relates to a medium additive for promoting protein expression and application thereof. Background technique [0002] Protein expression refers to a molecular biology technique that uses model organisms such as bacteria, yeast, animal cells or plant cells to express foreign gene proteins. It occupies a core position in genetic engineering technology. [0003] Mammalian expression systems have unique advantages in terms of protein initiation signals, processing, secretion, and glycosylation, and are suitable for expressing complete macromolecular proteins. The exogenous proteins produced by mammalian cells after translation and modification are far superior to prokaryotic expression systems and eukaryotic expression systems such as yeast and insect cells in terms of activity, and are closer to natural proteins, but they are complex in composition and require high operating t...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/10
CPCC12N5/0682C12N5/0686C12N2510/02C12N2500/38C12N2500/46C12N2500/32C12N2500/30
Inventor 赵峻岭谌颜鉄燕王静宏伟
Owner 内蒙古金源康生物工程股份有限公司
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