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From 2012.igem.org

Death Device

Outline

We plan to place endonuclease and YhjH gene downstream the xylR induce to make bacteria die and lose their adhesion when exposed to xylose.

Description

In order to terminate the overreaction of fatty acid metabolism and cellulose synthesis, we design an elaborate device to avoid it. Firstly, we determine to use endonuclease BglII(BBa_K112106, a protein which can kill the microbe without inducing cell lysis. Since AdrA but not YhjH contains this restriction site, our E.coslim will lose their adhesion more easily. To control the expression of endonuclease, a regulator is necessary. We select xylose as a controller because it is more healthy and hard to be absorbed by human. We find an ideal promoter BBa_K625002, which can be activated by the transcriptional regulator XylR. We hope that eating xylose will subsequently induce the expression of endonuclease, ending up with the death of our E.coslim. In addition, we use a pair of genes to strengthen the accuracy of controlling, namely YhjH. YhjH gene will counteract the impact of AdrA. YhjH protein can convert c-di-GMP in to GMP. Thus cells will lose their adhesion and become vulnerable to endonuclease.

Forestalling HGT

In human gut, there are billions of microbes. Horizontal gene transfer of those high efficient metabolic genes between GMOs and normal intestine residents may get out of control and cause disastrous effects. Therefore, a mechanism to prevent HGT is highly necessary. To prevent horizontal gene transfer, we designed a two plasmids system. In one plasmid, xlyR repressor encoding gene from Bacillus subtilis will be cloned. On the other plasmid, metabolic gene will be coupled with xylR repressed death system. In our GMOs, since death system is suppressed by xylR in low xylose concentration, the bacteria will not die. However, when the HGT of those high efficient metabolic genes happened, the death system will not be suppressed, and the recipient will be killed.