Maximizing Protein Expression

STEP® for recombinant proteins

For recombinant proteins, we have developed a plasmid–based expression technology that allows high product yield in CHO cells. Our STEP® plasmid contains a CMV promotor that drives the transcription of a single mRNA molecule from which two proteins are translated via an internal ribosomal entry site:

1. The protein of interest
2. A mutated, less potent selection marker protein (Zeocin)

The Zeocin marker mutant is combined with variable sizes of upstream open reading frames that variably hamper translation resulting in an adaptably stringent cell selection system. Because a cell needs to express high amounts of zeoR in order to survive selection, that cell, by default, will also produce high amounts of the protein of interest, as the gene of interest and the zeoR gene are expressed together on the single mRNA molecule. Three different zeoR mutants allow for different levels of stringency, depending on what is optimal for the protein of interest.

Proven Enhanced Expression Levels

To further enhance expression, two enhancer elements are cloned upstream and downstream of the expression cassette of the STEP® plasmid. The final STEP® plasmid, with minimum size enhancer elements and convenient multiple cloning sites for insertion of the gene of interest was completely synthesized under strict regulatory conditions to ensure suitability for developing pharmaceutical-grade cell lines.

STEP® technology has consistently proven to increase protein expression in CHO cells by generally more than 10-fold and taking about 12 weeks to generate stable cell clones. A highly stringent selection ensures our platform achieves extremely high protein expression levels in CHO cells without the need for gene amplification or time-consuming rounds of cell subcloning.