Team:Korea U Seoul/Project/Improvement

     Rice Guardian project was to build an engineered E. coli which detects and kills. However, our team had difficulties making plasmid construction which follows iGEM regulation. Therefore, our team decided to make Xanthomonas oryzae KACC10331 detecting E. coli beforehand. As we already discussed the mechanism of these bacteria, our team used red light emitting protein (RFP) as a final output instead of lysis and bacteriocin gene. Our alternative approach was successful and we decided to go one step further. Our team is conducting an experiment to add lysis and bacteriocin gene and this experiment will be done before iGEM championship.

     Addition of lysis and bacteriocin gene ensures death of nearby Xanthomonas oryzae KACC10331. Bacteriocins are proteinaceoustoxins produced by bacteria to inhibit the growth of similar or closely related bacterial strain(s). They are typically considered to be narrow spectrum antibiotics, though this has been debated . Lysis gene causes death of E. coli releasing bacteriocin inside the cell.

     Constitutive promoter will produce RaxR and RaxH. Protein synthesized from the genes allows cell to detect Ax21. When RaxR and RaxH proteins detect Ax21, They work as protein kinase (mechanism unknown) and stimulate bacteriocin and lysis gene. When lysis protein induces cell death of the E. coli , bacteriocin will burst into the surrounding and attacks Xanthomonas oryzae KACC10331.

     Designingthis project, our team’s final goal was to make standardized logic gate modulesfor organizing biological logic circuit. Therefore every designed logic gates’ inputsignal chemical had to be same with output signal chemical. (In this project,the chemical was name Chemical A ) And to show how wellthis works, we planned to make binary full adder. However due to the difficultyof finding Chemical A , experiment was not yet done. We recentlydecided use C4HSL for Chemical A . C4HSL is molecule usedin quorum sensing between bacteria, therefore it is density dependent. Thismeans the density of the bacteria in the logic gate effect on the signalintensity. We are still searching for the chemical substance, substitute forC4HSL.. While converting Chemical A to input signal, the original signal from the previous logic gatecan be either amplified or dampened. This directly effects on the logicoperation and change the total calculation result. To prevent this condition,further study of the conversion rate must be is needed.

     Because we designed the 2-digit binaryadder, currently we are not perfectly sure how linking more than one binaryadder module will effect on the calculation result. If we succeed in making thebinary adder module, experiment on 3 or more digit binary adder can be done asthe further study.