Transformation of Chlamydomonas reinhardtii CW-15 with the Hygromycin Phosphotransferase Gene as a Selectable Marker

To transform Chlamydomonas reinhardtiiDang. cells, plasmid pCTVHyg was constructed with the use of theEscherichia colihygromycin phosphotransferase gene (hpt) controlled by the SV40 early promoter. Cells of the CW-15 mutant strain were transformed by electroporation, with the yield reaching 10³ hygromycin-resistant (Hyg^R) clones per 10⁶ recipient cells. The exogenous DNA integrated in the Ch. reinhardtii nuclear genome showed stable transmission for approximately 350 cell generations, while hygromycin resistance was expressed as an unstable character. Codon usage was compared for the hptgene and Ch. reinhardtiinuclear genes. The results testified that codon usage bias, which is characteristic of Ch. reinhardtii, is not the major factor affecting foreign gene expression. The advantages of the selective system for studying Ch. reinhardtii transformation with heterologous genes are discussed.