Expression of human soluble TRAIL in Chlamydomonas reinhardtii chloroplast

The use of plants as bioreactors for the expression oftherapeutic proteins has developed into a new field in biotechnology research[1]. Plant systems provide simple genetic manipulation, low production costs, and low risk of contamination by animal viruse…

Expression of human soluble TRAIL in Chlamydomonas reinhardtii chloroplast

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) induces selectively apoptosis in various tumor cells and virus-infected cells, but rarely in normal cells. A chloroplast expression vector, p64TRAIL, containing the cDNA coding for the soluble TRAIL (sTRAIL), was constructed with clpP-trnL-petB-chlL-rpl23-rpl2 as Chlamydomonas reinhardtii plastid homologous recombinant fragments and spectinomycin-resistant aadA gene as a select marker. The plasmid p64TRAIL was transferred into the chloroplast genome of C. reinhardtii by the biolistic method. Three independently transformed lines were obtained by 100 mg/L spectinomycin selection. PCR amplification, Southern blot analysis of the sTRAIL coding region DNA and cultivation cells in the dark all showed that the exogenous DNA had been integrated into chloroplast genome of C. reinhardtii. Western blot analysis showed that human soluble TRAIL was expressed in C. reinhardtii chloroplast. The densitometric analysis of Western blot indicated that the expressed human sTRAIL protein in the chloroplasts of C. reinhardtii accounted for about 0.43%–0.67% of the total soluble proteins. These experimental results demonstrated the possibility of using transgenic chloroplasts of green alga as bioreactors for production of biopharmaceuticals.