Efficient expression of epidermal growth factor in Chlamydomonas reinhardtii CC400

Resumo

The application of the green alga Chlamydomonas reinhardtii as a bioreactor is not adequate because of the difficulties caused by efficiency expressing foreign genes. To improve this efficiency a plasmid containing the epidermal growth factor (EGF) gene and a bleomycin resistance gene (ble) was constructed. We amplified the EGF gene according to the codon usage of C. reinhardtii. The vector carrying 2 expression cassettes for EGF gene and ble gene was constructed by adding rbc promoter and rbc terminator. Transformants, selected on Tris-acetate-phosphate medium containing 15 mg/L bleomycin, were screened by PCR and confirmed by Southern blotting, which showed that 3 transgenic C. reinhardtii cells contained only one copy of EGF gene integrated in different 3 sites of C. reinhardtii CC400 genome. Then EGF protein content of 3 transformants was determined by EGF precoated ELISA, indicating that EGF gene was first expressed, although at a low level, in algal cells. The presented study, as an example for expressing heterologous gene in green alga, provided feasibility to improve the efficiency of transformation of C. reinhardtii.