An effective method to enhance transient gene expression by transfecting high-density CHO cells

Area of Science:

• Biotechnology
• Molecular Biology
• Cell Biology

Background:

• Chinese hamster ovary (CHO) cells are widely used for recombinant protein production.
• Transient gene expression (TGE) offers a faster alternative to stable gene expression (SGE) but often suffers from low protein yields.

Purpose of the Study:

• To optimize transient gene expression (TGE) systems in CHO cells for enhanced recombinant protein production.
• To improve protein yields in TGE by optimizing vector elements and transfection conditions.

Main Methods:

• Engineered vector regulatory elements (Kozak, polyA, introns, leader) and optimized transfection conditions for high-density suspension transfection.
• Utilized 40 kDa polyethylenimine (PEI) for transfection.
• Identified optimal DNA concentration (4 µg/mL), DNA:PEI mass ratio (1:5), and employed a repeated transfection strategy.

Main Results:

• Achieved a transfection efficiency of 76.34% with cell viability above 85%.
• Increased EGFP and SEAP levels by 5.33-fold and 4.13-fold, respectively.
• Enhanced production of therapeutic proteins, including recombinant type III human collagen (4.86-fold) and human serum albumin (5.50-fold).

Conclusions:

• High-density suspension transfection significantly boosts transient recombinant protein expression in CHO cells.
• The optimized approach improves transfection efficiency, expands the recombinant cell population, and maintains high cell viability.
• This method presents a promising strategy for high-yield transient transgene expression.