Part:BBa_K2350020
R0010-B0034-Holin-B0010-B0012-J23106-B0034-Endolysin-B0010-B0012
Holin-Endolysin
This mechanism which contains holin and endolysin was first proposed by 2014 Peking team. They used parts BBa_K1378031(holin) and BBa_K1378032(endolysin) for the suicide mechanism but we decided to choose other parts of holin and endolysin which labeled as BBa_K112000 and BBa_K112806. Moreover, we have also successfully construct the parts which containing both holin and endolysin.
In this composite part, holin functions as an important regulation of the whole suicide mechanism. Therefore, we choose a lactose-induced promoter, BBa_R0010. As for endolysin, we choose a constitutive promoter, BBa_J23106. In our design, we expect our suicide mechanism can reach maximize effect with this perfect combination.
Promoter, RBS, and Terminator
For more details of the connected promoter, RBS, and terminator, you can see in BBa_K2350018 for endolysin part and BBa_K2350019 for holin part.
Result
Figure 1 is restriction enzyme check electrophoresis result of holin-endolysin construct. The marker is 1kb. We use XmaI to digest the sample. The total length is 3843bp as expected.
Figure 1
Functional Test
The line graph below presents the general trend of O.D.600 value per hour after lactose is added.
As the figure shows, the suicide mechanism is induced immediately when lactose is added into the samples. Besides, we can see that the lactose concentration and the OD value of the bacterium with holin-endolysin construct are positively correlated, which means our suicide mechanism does work.
R0010-B0034-Holin-B0010-B0012-J23106-B0034-Endolysin-B0010-B0012
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 1035
Illegal NheI site found at 1058 - 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal AgeI site found at 1362
Illegal AgeI site found at 1432 - 1000COMPATIBLE WITH RFC[1000]
| None |