Melamine iteration
The point of our research is to help prove the concept of Programmed Evolution. We will try to do this by creating a system in E. coli that uses melamine deaminase to create ammeline. The presence of ammeline will be shown by using a riboswitch that will turn on a biosensor module and a fitness module. Our first goal is to create a riboswitch system that works well to show the presence of ammeline and then to work to build a system that efficiently produces ammeline from melamine.
Goals For Summer
1) create an effective riboswitch system
- retest riboswitches with biosensors (dissolve ammeline in DMSO, DEA, or NaOH)
- find best riboswitches, test them with the thyA fitness module (insert thyA riboswitch into mutant thyA- cells)
- use mathematical models to create new riboswitches
2) create an efficient melamine deanimase
- place melamine deaminase gene in the correct location in plasmid (MWSU)
- put both the gene and the riboswitch into the plasmid
- vary the promoter, RBS, and compy number to get efficient ammeline production
Eckdahl's riboswitches File:Ammeline Riboswitch Sequences.docx
Ammeline Riboswitch Numbering System
1. AddA
2. M6
3. M6C
4. M6’
5. M6’’
6. M6C’
7. M6C’’
8. M6C (A172G Mutation)
9. M6C (T92C Mutation)
10. M6C’ (T167C Mutation)
- note - change to RNA before using
