Part:BBa_K302018
Subtilin immunity
Subtilin is a lantibiotic peptide produced naturally by Bacillus subtilis ATCC 6633 but not by Bacillus subtilis 168.
This part encodes two separate subtilin immunity systems — a membrane bound protein that binds subtilin and prevents it from penetrating the cell membrane to form pores, and a transporter which removes subtilin from the membrane — which together provide a high degree of subtilin resistance.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal BglII site found at 837
Illegal BglII site found at 1585 - 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal NgoMIV site found at 1852
- 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI.rc site found at 1132
Illegal SapI site found at 1412
Illegal SapI.rc site found at 2135
User Reviews
Liaoliao_Gao
In 2022, ZJU-China iGEM team has characterized the output of this part in a novel chassis E. coli BL21(DE3). The result was documented in the experience page and the main page of (BBa_K302018).
Contribution from iGEM2022 ZJU-China
The sequence of (BBa_K302018) was synthesized and cloned into the PHY-plasmid to obtain the recombinant expression vector
We first tested the inhibitory concentration of subtilisin by zone of inhibition experiment, using BL21 transformed by PHY-Immunity plasmid or PHY-Empty plasmid as control. The concentration of subtilisin were 100mg/ml, 50mg/ml, 25mg/ml, 12.5mg/ml.
Then, we prepared normal LB culture plates with different concentration of subtilisin, including 0.1mg/ml, 0.2mg/ml, 1mg/ml and 10mg/ml, in individual dishes. Then we inoculated BL21 transformed by immunity plasmid or PHY-empty plasmid as control in the plates. For the immunity group, some colonies grew on the surface of the media with 1mg/ml subtilisin, while the control group didn’t. This suggested that the PHY-Immunity plasmid worked well in BL21.
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