Part:BBa_K4849014:Design

MtrB - periplasmic decaheme cytochrome protein

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  COMPATIBLE WITH RFC[1000]

Design Notes

Gel Showing MtrB extracted by PCR:

 

 

 

Gene Sequence: ATGAAATTTAAACTCAATTTGATCACTCTAGCGTTATTAGCCAACACAGGCTTGGCCGTCGCTGCTGATGGTTATGGTCTAGCGAATGCCAATACTGAAAAAGTGAAATTATCCGCATGGAGCTGTAAAGGCTGCGTCGTTGAAACGGGCACATCAGGCACTGTGGGTGTCGGTGTCGGTTATAACAGCGAAGAGGATATTCGCTCTGCCAATGCCTTTGGTACATCCAATGAAGTGGCGGGTAAATTTGATGCCGATTTAAACTTTAAAGGTGAAAAGGGTTATCGTGCCAGTGTTGATGCTTATCAACTCGGTATGGATGGCGGTCGCTTAGATGTCAATGCGGGCAAACAAGGCCAGTACAACGTCAATGTGAACTATCGCCAAATTGCTACCTACGACAGCAATAGCGCCCTATCGCCCTACGCGGGTATTGGTGGCAATAACCTCACGTTACCGGATAACTGGATAACAGCAGGTTCAAGCAACCAAATGCCACTCTTGATGGACAGCCTCAATGCCCTCGAACTCTCACTTAAACGTGAGCGCACGGGGTTGGGATTTGAATATCAAGGTGAATCCCTGTGGAGCACCTATGTTAACTACATGCGTGAAGAGAAAACCGGCTTAAAACAAGCCTCTGGTAGCTTCTTCAACCAATCGATGATGTTAGCAGAGCCGGTGGATTACACCACTGACACCATTGAAGCGGGTGTCAAACTCAAGGGTGATCGTTGGTTTACCGCACTCAGTTACAATGGGTCAATATTCAAAAACGAATACAACCAATTGGACTTTGAAAATGCTTTTAACCCCACCTTTGGTGCTCAAACCCAAGGTACGATGGCACTCGATCCGGATAACCAGTCACACACCGTGTCGCTGATGGGACAGTACAACGATGGCAGCAACGCACTGTCGGGTCGTATTCTGACCGGACAAATGAGCCAAGATCAGGCGTTAGTGACGGATAACTACCGTTATGCTAATCAGCTCAATACCGATGCCGTCGATGCCAAAGTCGATCTACTGGGTATGAACCTGAAAGTCGTTAGCAAAGTGAGCAATGATCTTCGCTTAACAGGTAGTTACGATTATTACGACCGTGACAATAATACCCAAGTAGAAGAATGGACTCAGATCAGCATCAACAATGTCAACGGTAAGGTGGCTTATAACACCCCTTACGATAATCGTACGCAACGCTTTAAAGTTGCCGCAGATTATCGCATTACCCGCGATATCAAACTCGATGGTGGTTATGACTTCAAACGTGACCAACGTGATTATCAAGACCGTGAAACCACGGATGAAAATACCGTTTGGGCCCGTTTACGTGTAAACAGCTTCGATACTTGGGACATGTGGGTAAAAGGCAGTTACGGTAACCGTGACGGCTCACAATACCAAGCGTCTGAATGGACCTCTTCTGAAACCAACAGCCTGTTACGTAAGTACAATCTGGCTGACCGTGACAGAACTCAAGTCGAAGCACGGATCACCCATTCGCCATTAGAAAGCCTGACTATCGATGTTGGTGCCCGTTACGCGTTAGATGATTATACCGATACTGTGATTGGATTAACTGAGTCAAAAGACACCAGTTATGATGCCAACATCAGTTATATGATCACCGCTGACTTACTGGCAACCGCCTTCTACAATTACCAAACCATTGAGTCTGAACAGGCGGGTAGCAGCAATTACAGCACCCCAACGTGGACAGGCTTTATAGAAGATCAGGTAGATGTGGTCGGTGCAGGTATCAGCTACAACAATCTGCTGGAGAACAAGTTACGCCTAGGACTGGACTACACCTATTCCAACTCCGACAGTAACACTCAAGTCAGACAAGGTATCACTGGCGACTATGGTGATTATTTTGCCAAAGTGCATAACATTAACTTATACGCTCAATATCAAGCCACCGAGAAACTCGCGCTGCGCTTCGATTACAAAATTGAGAACTATAAGGACAATGACGCCGCAAATGATATCGCCGTTGATGGCATTTGGAACGTCGTAGGTTTTGGTAGTAACAGCCATGACTACACCGCACAAATGCTGATGCTGAGCATGAGTTACAAACTCTAA

 

We extracted the Gene from Shewanella oneidensis MR-1 (NCBI:txid211586) genomci DNA using Primers that introduced cynogate assmbly system overhangs (refer IGEM Edinburgh 2023 wiki)

 

MtrB Forward Primer: CAGTGAAGACAtAATGAAATTTAAACTCAATTTGATCAC

MtrB Reverse Primer: CAGTgaagacATaagcTTAGAGTTTGTAACTCATGCTC

 

The overhangs were designed to be cut by BbsI - this would allow introduction into the Cynogate Lv0 acceptor vector

Source

Shewanella oneidensis MR-1 (NCBI:txid211586)

References

1. Ross, Daniel E., et al. "Characterization of protein-protein interactions involved in iron reduction by Shewanella oneidensis MR-1." Applied and environmental microbiology 73.18 (2007): 5797-5808.

2. Breuer, Marian, et al. "Multi-haem cytochromes in Shewanella oneidensis MR-1: structures, functions and opportunities." Journal of The Royal Society Interface 12.102 (2015): 20141117.