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Revision as of 02:48, 13 December 2017
Description
Light-activated gene expression
Light is an attractive mode of gene regulation that provides high spatio-temporal resolution with relatively low levels of toxicity. In order to add to the genetic toolbox, we characterized a novel light-activated gene regulation system that combines the DNA-binding region of LacI with the light inducible LOV (Light Oxygen Voltage) domain. The characterization assay was performed by measuring the fluorescence output of a LacILOV-regulated reporter under blue light illumination. We then computationally modelled the structure of our protein and identified key mutations to optimize its activity.
Identifying and informing stakeholders
In order to inform future applications of our tool, we identified key stakeholders that would be impacted by potential uses of LacILOV including experts, businesses, the public and future scientists. To this end, we developed resources to promote interest and meaningful interdisciplinary dialogue between researchers and the public. This was achieved through a podcast series exploring the interaction of synthetic biology with different disciplines, a synthetic biology workshop for burgeoning scientists and an iconathon to promote collaborations between scientists and artists.
Applying LacILOV to human gene editing
However, in order to demonstrate the utility of our tool, we decided to apply LacILOV to a specific area of study. As a rapidly evolving field with a clear need for flexible and stringent gene regulation, we focused our efforts on human gene editing. We designed and modelled a light activated switch to control CRISPR-Cas9 activity by putting guide RNAs and anti-CRISPR proteins under LacILOV-regulated promoters. We then investigated the ethical and technical concerns of our stakeholders through an interview series involving scientists, engineers, physicians, advocacy groups and religious leaders. We integrated their comments into our design in two key ways. Firstly, we addressed technical concerns about the use of light regulation in human gene editing by developing hardware to pave the way for our system to be validated in the stem cell cultures. Secondly, based on the different opinions on the ethical applications of human gene editing, we identified the need for clear ethical guidelines. Using the recommendations of the 2017 report on Human Gene Editing: Science, Ethics and Governance by the National Academy of Science, we developed a guide and evaluated our efforts and potential improvements.
