Codon optimization

EGF (synthetic gene) CaMV 35S promoter/ nos terminator Codon-optimized for high level peptide expression in Escherichia coli N. tabacum (leaves) 0.001% ofTSP 38... 

Fig. 8.3 Expression of IGF-1 in transgenic chloroplasts after continuous light exposure for 13 days. IGF-1 expression is shown as a percentage of the total soluble protein. IGF-1 n is the native gene and IGF-1 s is the chloroplast codon-optimized gene.

GFP has also been proposed as a successor to the Ames and SOS chromotest. Billinton et al. [8] obtained a reporter system, employed as genotoxicity biosensor, that uses eukaryotic cells (the baker yeast Saccharomyces cerevisiae) instead of bacteria. The strain produces green fluorescent protein, codon optimized for yeast, when DNA damage has occurred. It was demonstrated that the reporter does not falsely respond to chemicals that delay mitosis, and responds appropriately to the genetic regulation of DNA repair. 

Nagata, T., Uchijima, M., Yoshida, A., Kawashima, M. and Koide, Y. (1999) Codon optimization effect on translational efficiency of DNA vaccine in mammalian cells analysis of plasmid DNA encoding a CTL epitope derived from microorganisms. Biochem. Biophys. Res. Commun., 261, 445—451. 

Kim CH, Oh Y, Lee TH (1997), Codon optimization for high-level expression of human erythropoietin (EPO) in mammalian cells - human or yeast codon usage effect on over-expression in 293T cell culture, Gene 199 293-301. 

Chen et al. (2008) have expressed a codon-optimized form of the CM4 peptide in E. coli. In this case, two alternative expression/purification systems were examined namely the widely used glutathione-S-transferase (GST) and a chitin-binding domain (CBD) system with associated intein splicing (New England Biolabs Inc.). The GST system failed to allow recovery of expressed fusion protein. In contrast, the CBD/intein system allowed the recovery of 110 mg/L fusion protein with a final RP-HPLC purification step yielding 2.1 mg/L of pure peptide which displayed antimicrobial activity against E. coli Ki2D3i and Salmonella. 

Burgess-Brown NA, Sharma S, SobottF, Loenarz C, Oppermann U, Gileadi O (2008) Codon optimization can improve expression of human genes in Escherichia coli a multi-gene study. Protein Expr Purif 59 94-102... 

Fuglsang A (2003) Codon optimizer a freeware tool for codon optimization. Protein Expr Purif 31 247-249... 

Heterologous expression of synthesized genes, involving codon optimization of the external genetic information to achieve an adequate translational quantity of the recombinant protein (91) (Fig. IH). 

Because HIV-1 is a human pathogen, there is some concern about the use of HIV-1-based lentiviral vectors. The current HIV-1 lentiviral vector system is deprived of all accessory proteins (except rev) and viral sequences in the vectors have been minimized therefore, replication of these vectors is highly disabled and the possibility of homologous recombination is minimized. In addition, codon-optimization of the packaging construct further decreases the risk of homologous recombination. Furthermore, changes in the codons makes the production of the structural proteins independent of rev, and therefore additional viral sequences (RRE) can be eliminated from the packaging construct (82). 

Kotsopoulou E, Kim VN, Kingsman AJ, Kingsman SM, Mitrophanous KA. 2000. A Rev-independent human immunodeficiency virus type 1 (HlV-l)-based vector that exploits a codon-optimized HIV-1 gag-pol gene. J. Virol. 74 4839-52... 

Yeast. Two yeast stress promoters RAD54 and RNR2 were fused transcriptionally to GFP to screen for general stress (Afanassiev et al., 2000). Billinton et al. (1998) developed a codon-optimized GFP version for yeast. 

Yazdani, S. S., Shakri, A. R., Pattnaik, P., Rizvi, M. M., and Chitnis, C. E. (2006). Improvement in yield and purity of a recombinant malaria vaccine candidate based on the receptorbinding domain of Plasmodium vivax Duffy binding protein by codon optimization. Biotechnol. Lett. 28,1109-1114. 

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