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Effects of the dilution rate on cell cycle distribution and PEI-mediated transient gene expression by CHO cells in continuous culture

Paper ID Volume ID Publish Year Pages File Format Full-Text
34554 45033 2013 9 PDF Available
Title
Effects of the dilution rate on cell cycle distribution and PEI-mediated transient gene expression by CHO cells in continuous culture
Abstract

•Recombinant prothrombin was produced by suspension transient transfection.•A novel PEI polymer-PEI MAX was chosen as a superior gene delivery vehicle.•Continuous culture provides better physiological state for TGE than batch culture.•Transient transfection is greatly affected by cell cycle distribution.•Mitosis is essential for transient gene expression.

In this study, a continuous culture system was applied to mammalian cells on large scale, and polyethyleneimine (PEI) mediated transient gene expression (TGE). PEI MAX 40,000 was chosen as a superior reagent from three types of PEI. The cell cycle distribution of cells in batch and continuous cultures was determined, in which the effects of cell cycle distribution on transfection efficiency, post-transfection proliferation and recombinant prothrombin expression were evaluated. Compared with cells from end-log and plateau phase in batch culture, cells from mid-log phase possessed a larger fraction of S and G2/M phase cells and a smaller fraction of G1 phase cells. In the continuous culture, the fraction of cells in the S and G2/M phases increased and the fraction of cells in the G1/G0 phase decreased with increasing dilution rates. Cells from the continuous culture run at highest dilution rate had excellent proliferation, transfection efficiency and protein expression. These results were confirmed by transfecting cells synchronized to different phases. The G2/M arrested cells exhibited a nearly 10-fold increase in recombinant human prothrombin production relative to that of non-dividing cells. The use of continuous culture for large scale transfection demonstrated a better cell physiological state for TGE process.

Keywords
Cell cycle distribution; Transient gene expression; Transient transfection; Recombinant protein; Cell cycle synchronization; Continuous culture
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Effects of the dilution rate on cell cycle distribution and PEI-mediated transient gene expression by CHO cells in continuous culture
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Publisher
Database: Elsevier - ScienceDirect
Journal: Process Biochemistry - Volume 48, Issue 10, October 2013, Pages 1572–1580
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us