Team:Hong Kong HKU/Description/ex
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Concerns about accidental leakage of GMO
Advancement in science and engineering has led us to the era of synthetic biology. By reprogramming living things at DNA level, synthetic biologists could introduce new characteristics or modify existing properties to create synthetic organisms for the sake of human well-beings. For example, bacteria and yeast have been engineered for drug production that can hardly be obtained in nature1 and for cleaning-up large scale environmental pollution2. Given the rapid development of synthetic biology, more sophisticated organisms with de novo applications may be created in the future. Meanwhile the expanding applications of synthetic biology have also raised public concerns over the risk of accidental leakage of synthetic organisms which may jeopardize the ecosystem and public health3,4.
Control of leakage
Although no one has fully understood the actual impacts of escaped synthetic organisms on the environment yet, various containment policies have been adopted to minimize accidental leakage of genetically modified organisms (GMOs). Physical containment, such as proper handling of genetically modified microorganisms (GMM) with suitable containment equipment and implementation of laboratory practice is one of the simplest way to prevent escape of synthetic organisms3. However this strategy may not be applicable to open-setting applications, for example biomining5 and biosensing6 which often involve deliberate dispersal of GMO covering large area.
To address this problem, intrinsic containment or ‘built-in safeguard’ have been developed to control unintentional release of GMM. One of them is to design auxotrophic organisms by removing metabolic pathways responsible for synthesizing essential nutrients, such that modified organisms can only survive in the environment with addition of essential nutrients7. Alternatively, lethality can be induced by built-in killswitch which will be activated when GMM escape from specific environment. For instance, iGEM13_BGU_Israel has designed an IPTG-and-arabinose-regulated repressible killswitch using holin and lysozyme, in which escape from medium containing IPTG or arabinose would lead to lysis of bacterial cell wall, and hence cell death.
Toxin-based killswitches seem to be effective in eliminating escaped GMM but undestroyed plasmid and chromosomal DNA could remain intact in the environment for months8. Moreover, a recent astrobiological research has demonstrated that up to 35% of plasmid DNA remain functional even after expose to high temperature and pressure9. Chances are that recombinant DNA may be taken up by natural occurring bacteria even though escaped synthetic organisms are destroyed. Thus, in order to fully unleash the potential of synthetic biology, it is necessary to develop more comprehensive containment strategies to control the leakage of GMO and recombinant DNA.
Exploring new containment device
In light of the demand of better containment strategies, we would like to explore the potential of CRISPR/Cas system as an intrinsic containment device. In this project, we are going to design two prototype repressible killswitches based on two bacteria immune systems against viral infection – CRISPR/Cas3 (originated from E. coli MG1655) and CRISPR/Cas9 (originated from S. pyogenes SF370). In addition, we would like to compare the killing efficiencies of the devices and test our devices out in a simulated scenario using E. coli BL21 (DE3) as the chassis.
For the detail of our containment device please visit our Design page.