Team:Aberdeen Scotland/WetLab/lacilatch

Introduction:

The LacI latch module was designed to maintain the system in the ON state once it has been triggered, by repressing LacI production, and to induce timed cell lysis by repressing TetR production. These two functions enable production of our target proteins even after IPTG is no longer present, thus producing Elastin for a certain time period after a leak has been closed, allowing for extra strengthening of the weak spot.

The LacI latch module consists of a promoter with a CI operator, a Tet Repressor gene and the LacI gene. The TetOperon and the Lysis Cassete which was developed by the 2008 Berkley iGEM team is then added to the module.

How it works:

Please note that for the purpose of simplicity Quorum sensing is ignored in this section. The system would not turn on with just the IPTG stimuli.

The normal state of our system is illustrated in the picture above. LacI is expressed and binds to the Lac Operator thus repressing expression of our glue and the CI repressor. At the same time TetR is expressed and and blocks expression of Holin, and Endolysin. Note that Antiholin is not affected by this and is in fact expressed since it is under control of a separate constitutive promoter.

Once IPTG is induced our target Gene starts being expressed in addition to this the CI repressor gene can be expressed.

Once the CI repressor is synthesised it binds to the CI operon, this halts expression of LacI and TetR. LacI and TetR levels then start to fall inside of the cell since no more molecules are being made. At this point TetR is still repressing Holin and Endolysin, which creates a time delay before Holin and Endolysin are expressed and the cell lysis.

Once TetR and LacI have completely degraded, the system is latched on, and it will carry on expressing our product until it lysis.

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