Team:RHIT/Design
Design
This project employed the use of a new arsenic metallothionein (BBa_K3275000), to test its capabilities to bioremediate arsenite ions from aqueous solutions. To this end, two plasmid constructs were designed, denoted A1 and A2. A1 employs the use of an ArsR operon (BBa_J33201) to control the transcription of arsenic metallothionein based on the amount of arsenite in the cell. A double terminator (BBa_B0017) ends the sequence. A2 on the other hand, is run by a constitutive promoter (BBa_J23100), and a strong RBS (BBa_B0030). The metallothionein protein produced by our part can bind to six total arsenite ions due to its twenty total cysteins.
iGEM continues to partner with IDT to supply teams with free DNA synthesis, in the form of GBlocks. Each part that was synthesized came in the format: iGEM prefix, promoter/ArsR, RBS, metallothionein, iGEM suffix. This enabled the same primers to be used in all PCR, and enabled easy digestion and ligation.
Multiple duet vectors were examined for use as the backbone in our characterization tests this summer. pETDuet-1 was selected for use, due to its very close EcoR1 and Pst1 cut sites, both to each other and to a promoter, RBS, and start codon. The part we tested was a SOD gene (BBa_K847004) intended for use against, in part, copper.
Our Plasmids
