Codon optimization of genes for efficient protein expression in mammalian cells by selection of only preferred human codons

•A simple method of codon optimization for expressing in mammalian cells.•Our method was named the “preferred human codon-optimized method.”•Genes were synthesized with only preferentially used human codons.•The GC content of preferred human codon-optimized genes was over 60%.•Preferred human codon-optimized genes were efficiently expressed in mammalian cells.

A simple design method for codon optimization of genes to express a heterologous protein in mammalian cells is described. Codon optimization was performed by choosing only codons preferentially used in humans and with over 60% GC content, and the method was named the “preferred human codon-optimized method.” To test our simple rule for codon optimization, the preferred human codon-optimized genes for six proteins containing photoproteins (aequorin and clytin II) and luciferases (Gaussia luciferase, Renilla luciferase, and firefly luciferases from Photinus pyralis and Luciola cruciata) were chemically synthesized and transiently expressed in Chinese hamster ovary-K1 cells. All preferred human codon-optimized genes showed higher luminescence activity than the corresponding wild-type genes. Our simple design method could be used to improve protein expression in mammalian cells efficiently.