Difference between revisions of "Part:BBa K302018"
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<partinfo>BBa_K302018 parameters</partinfo> | <partinfo>BBa_K302018 parameters</partinfo> | ||
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| + | ==User Reviews== | ||
| + | Liaoliao_Gao<br> | ||
| + | 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 (<partinfo>BBa_K302018</partinfo>).<br> | ||
| + | ===Contribution from iGEM2022 ZJU-China=== | ||
| + | <p>The sequence of (<partinfo>BBa_K302018</partinfo>) was synthesized and cloned into the PHY-plasmid to obtain the recombinant expression vector </p> | ||
| + | <p>We first tested the inhibitory concentration of subtilisin by <b>zone of inhibition experiment</b>, 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. </p> | ||
| + | <div align="center">[[File:GLL-2.png|600px]]</div> | ||
| + | <center>Fig1 Zone of inhibition experiment for subtilisin in BL21. | ||
| + | </center> | ||
| + | <!-- --> | ||
| + | <p>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. | ||
| + | <p><b>Results：</b>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.</p> | ||
| + | <div align="center">[[File:GLL-1.png|600px]]</div> | ||
| + | <center>Fig2 The colonies growth condition in plates with different concentration of subtilisin | ||
Revision as of 03:50, 26 September 2022
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. <p>Results：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.
