Coding

Part:BBa_K2997001

Designed by: Ting-Han Sung   Group: iGEM19_NCKU_Tainan   (2019-10-12)
Revision as of 18:50, 19 October 2019 by Tanya0809 (Talk | contribs) (→‎Reference)


Bacteriocin full length (CBM-B)

In the human gut microbiome, Clostridium related species have been reported to have the highest conversion capability of fermenting tyrosine into p-cresol[1]. To target the root of p-cresol accumulation, reducing the population of Clostridium is needed. We decide to use C. difficile as a model of p-cresol producing bacteria because it is a popular research target due to its notorious infectious ability.

Luckily, iGEM NCKU 2019 was kindly supported by one of our PIs, Professor I-Hsiu, an assistant professor from the Microbiology and Immunology Department, who is currently devoting himself in the field of developing novel therapeutic approach for C. difficile infection. He kindly provided us with a plasmid containing a bacteriocin gene (CBM-B) from Clostridium Butyricum Miyairi. This bacteriocin was proven to have bactericidal activity against certain strains of Clostridium, including C. difficile[2].

As proof of concept that bacteriocin is able to inhibit Clostridium growth, we did a spot-on-lawn assay using purified bacteriocin protein provided by advisors to observe the inhibition zone formation. As shown in Fig.1 below, a clear inhibition zone formed in the middle of the BHI plate streak with C. difficile R20291 strain.



Fig. 1. Spot-on-lawn test using 5 μl purified bacteriocin, inhibition zone formation in the middle of the plate can clearly be seen.

Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal SpeI site found at 396
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal SpeI site found at 396
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal SpeI site found at 396
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal SpeI site found at 396
  • 1000
    COMPATIBLE WITH RFC[1000]


Reference

[1] Saito, Y., Sato, T., Nomoto, K., & Tsuji, H. (2018). Identification of phenol- and p-cresol-producing intestinal bacteria by using media supplemented with tyrosine and its metabolites. FEMS Microbiology Ecology, 94(9).

[2] Nakanishi, S., & Tanaka, M. (2010). Sequence analysis of a bacteriocinogenic plasmid of Clostridium butyricum and expression of the bacteriocin gene in Escherichia coli. Anaerobe, 16(3), 253–257

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