Bioengineering for Cleaner Water: iGEM at ASU

Arsenic contamination in groundwater is a serious problem both in local Arizonan communities and abroad: prolonged exposure to arsenic contamination can cause cancer, vascular damage, and liver failure. This project aims to engineer the microalgae Chlamydomonas reinhardtii to sequester arsenic out of water. Metallothionein, arsenate reductase, and ferritin were integrated into the microalgae via the pASapI plasmid. The plasmid rescues function of the photosystem II gene, leveraging the ability to photosynthesize as a selective trait. Metallothionein and ferritin bind the two most common forms of arsenic: arsenite and arsenate, respectively. Arsenate reductase catalyzes the reduction of arsenate to arsenite, allowing for the ultimate sequestration of the toxic metal to occur in the chloroplast. The algae was transformed using a biolistic device, to create three mutant strains, expressing Metallothionein (MT), Arsenate Reductase (ArsC)-HA, and MT-6xHIS plasmids respectively. When testing the fluorescence output of these three strains, they showed a maximum quantum yield of photosystem II comparable to that of the wildtype algae, indicating that the rescue gene had been incorporated into the chloroplast genome properly. Strains were exposed to arsenic-containing media at 50ppb and 500 ppb for 48 and 72 hours to determine the arsenic sequestration rate. Arsenic concentration in the supernatant was measured using ICP-MS analysis and sequestration rate was calculated in terms of arsenic concentration per fold growth of algae. The normalized arsenic sequestration rates of tagged protein expressing strains at 50 ppb were significantly higher than wildtype.