Abstract
Three transfection reagents, Lipofectamine® 2000, TransIT-PRO® and linear 25 kDa polyethylenimine were evaluated for transient expression of enhanced green fluorescent protein in Chinese hamster ovary cells. TransIT-PRO® was found to be more efficient under the examined conditions, but comes at an increased cost compared to the widely used PEI.
Highlights
Transient gene expression (TGE) is capable of producing recombinant proteins faster and cheaper than the standard stable cell line development process, making it an attractive alternative for the provision of material for preclinical trials and toxicological studies [1]
From the results (Figure S1), we concluded that a 1:2 DNA:Lipofectamine® 2000 ratio gave a better distribution of enhanced green fluorescent protein (EGFP)-C3 expression, with an exponential increase of EGFP fluorescence and a maximum protein expression at 200 hours (Figure S1(a))
In the case of PEI (Figure S1(b)), a 1:4 DNA:PEI ratio resulted in the highest levels of fluorescence, while DNA:PEI ratios above 1:5 showed very low protein expression levels. This is in contrast with Xie et al [9] who suggested that a higher PEI concentration assisted DNA delivery into the cell nucleus and recommended a 1:5 ratio for an EGFP vector transfected into Chinese hamster ovary (CHO)-DG44 cells
Summary
Transient gene expression (TGE) is capable of producing recombinant proteins (rProteins) faster and cheaper than the standard stable cell line development process, making it an attractive alternative for the provision of material for preclinical trials and toxicological studies [1]. PEI works by forming polyplexes with target DNA These positively charged polyplexes interact with the negatively charged host cell membrane and release DNA into the cell. TransIT-PRO® employs both mechanisms of action and works by forming lipopolyplexes with DNA, which forms electrostatic interactions with the negatively charged cell membrane. These lipopolyplexes are believed to be taken into the cell via endocytosis. The use of this reagent involves an optional proprietary transfection booster PRO Boost, which was shown to enhance gene expression in a medium-dependent manner
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