And an oxygen sensitive reporter system Motivation Microbial Fuel - PowerPoint PPT Presentation

And an oxygen sensitive reporter system

Motivation

Microbial Fuel Cells Electricity Produced Bacteria Incoming ½ O 2 Incoming O 2 wastewater ` Outgoing H 2 O Outgoing CO 2 Anode Cathode Semi-permeable membrane

Genetic design NirB RBS LOV Terminator

4Fe-4S O 2 Ribosome mRNA } FNR FNR Dimerization FNR FNR Translation Transcription RNA How does our openon works? Polymerase FNR= Ferredoxin-NADP+ reductase

The LOV Domain Optimized LOV for Wild type LOV gene from Shewanella oneidensis Bacillus subtilis Removal Cysteine amino acid codon Adding TAA stop codon Added Alanine codon

Safety • Sterilizing Work Stations • Personal Safety • Equipment testing • Proper disposal of waste materials • Proper supervision

Methods Design Synthesis

Transformations Transformation pSB1C3/RFP pSB1C3/LOV

Electrophoresis pSB1C3/NirB pSB1C3/LOV 4 Kb 2 Kb

Sequencing CLUSTAL 2.1 multiple sequence alignment LOV --------------------------------------------GAATTC 14 02-027-08-VF2_D11.ab1 TTCNGATAAAAAAAATCCTTAGCTTTCGCNANNGNNGATTTCTGGAATTC 100 02-027-06-VF2_C11.ab1 TTCAGATAAAAAAAATCCTTAGCTTTCGCNNAGGANGATTTCTGGAATTC 99 ****** LOV GCGGCCGCTTCTAGAGATGGCTTCTTTCCAATCTTTCGGTATCCCAGGTC 64 02-027-08-VF2_D11.ab1 GCGGCCGCTTCTAGAGATGGCTTCTTTCCAATCTTTCGGTATCCCAGGTC 150 02-027-06-VF2_C11.ab1 GCGGCCGCTTCTAGAGATGGCTTCTTTCCAATCTTTCGGTATCCCAGGTC 149 ************************************************** LOV AACTGGAAGTTATCAAAAAAGCTCTGGATCACGTTCGTGTTGGTGTTGTT 114 02-027-08-VF2_D11.ab1 AACTGGAAGTTATCAAAAAAGCTCTGGATCACGTTCGTGTTGGTGTTGTT 200 02-027-06-VF2_C11.ab1 AACTGGAAGTTATCAAAAAAGCTCTGGATCACGTTCGTGTTGGTGTTGTT 199 ************************************************** LOV ATCACTGATCCAGCTCTGGAAGATAACCCAATCGTTTACGTTAACCAAGG 164 02-027-08-VF2_D11.ab1 ATCACTGATCCAGCTCTGGAAGATAACCCAATCGTTTACGTTAACCAAGG 250 02-027-06-VF2_C11.ab1 ATCACTGATCCAGCTCTGGAAGATAACCCAATCGTTTACGTTAACCAAGG 249 ************************************************** LOV TTTCGTTCAAATGACTGGTTACGAAACTGAAGAAATCCTGGGTAAAAACG 214 02-027-08-VF2_D11.ab1 TTTCGTTCAAATGACTGGTTACGAAACTGAAGAAATCCTGGGTAAAAACG 300 02-027-06-VF2_C11.ab1 TTTCGTTCAAATGACTGGTTACGAAACTGAAGAAATCCTGGGTAAAAACG 299 ************************************************** LOV CTCGTTTCCTGCAAGGTAAACACACTGATCCAGCTGAAGTTGATAACATC 264 02-027-08-VF2_D11.ab1 CTCGTTTCCTGCAAGGTAAACACACTGATCCAGCTGAAGTTGATAACATC 350 02-027-06-VF2_C11.ab1 CTCGTTTCCTGCAAGGTAAACACACTGATCCAGCTGAAGTTGATAACATC 349 ************************************************** LOV CGTACTGCTCTGCAAAACAAAGAACCAGTTACTGTTCAAATCCAAAACTA 314 02-027-08-VF2_D11.ab1 CGTACTGCTCTGCAAAACAAAGAACCAGTTACTGTTCAAATCCAAAACTA 400 02-027-06-VF2_C11.ab1 CGTACTGCTCTGCAAAACAAAGAACCAGTTACTGTTCAAATCCAAAACTA 399 ************************************************** LOV CAAAAAAGATGGTACTATGTTCTGGAACGAACTGAACATCGATCCAATGG 364 02-027-08-VF2_D11.ab1 CAAAAAAGATGGTACTATGTTCTGGAACGAACTGAACATCGATCCAATGG 450 02-027-06-VF2_C11.ab1 CAAAAAAGATGGTACTATGTTCTGGAACGAACTGAACATCGATCCAATGG 449 **************************************************

Human Practices Waste Water CO2 Anode Cathode

Where do we go from here? ● Retry NirB Plasmid formation ● Finish 3A assembly with LOV and NirB ● Utilizing the microbial fuel cell with Schwenella

Acknowledgements