Glycerol Stock Preparation

1. Inoculated TSB with MR-1 (either from plate or glycerol stock) and grow overnight.
2. add glycerol to culture to final concentration of 80% glycerol.
3. place 1ml in 1.5ml freezer safe eppendorf tubes (do not fill eppendorfs with more than 1.5ml to allow for expansion by freezing).

Microbual Fuel Cell Assembly

1. cut two 3 inch PVC tubes .
2. sandwich them with nafion cation exchange membrane between them.
3. add caulking around both sides of membrane-tube junction.
4. let dry for 15 minutes.
5. repeat step 3 and 4.
6. cut off excess membrane around junction.
7. add one more layer to seal up junction.
8. let dry overnight.
9. cut 2.5in-3in x 12in of carbon cloth
10. roll around tip of finger.
11. weave titanium wire close to end edge of roll.
12. fold titanium wire up the length of the roll.
13. twist excess wire around itself.
14. cut wire at .5in-1in.
15. push through stopper.
16. seal hole from wire with caulking.
17. let dry overnight. (two of these make up the cathode and the anode)
18. pour potassium ferricyanide in one chamber of the cell.
19. carefully push stopper with carbon cloth to plug the chamber.
20. push hollow needle through stopper to let air out of chamber.
21. pour overnight culture of MR-1 into other chamber.
22. place glass slide in chamber.
23. carefully push stopper with carbon cloth to plug the chamber.
24. push new sterile hollow needle in stopper into air bubble in chamber to let out any CO2 bacteria will produce.
25. open ExcelINX up on computer.
26. connect electrodes from DMM to cathode and anode (remember which channels are being used).
27. start ExcelINX.

Plasmid Prep