The following was authored by iGEM team member Ciara McCarthy and is reposted with permission from the Cambridge Consultants blog.
Whilst the majority of students had long since escaped the labs and libraries of Cambridge for the summer, 10 intrepid students remained in the Plant Sciences department. We were a mixture of engineers and natural scientists taking part in iGEM – an international synthetic biology competition. Teams from universities across the world participate, building genetically engineered systems that aim to have ‘a positive impact on their communities and the world’.
This year, the aim of the Cambridge team was to create a toolkit for algal chloroplast engineering, which is a process that holds great potential for producing everything from biofuels to edible vaccines efficiently and in large quantities.
However, chloroplast engineering has its drawbacks. Chloroplasts contain many copies of the chloroplast genome – for example, the chloroplasts of the algae that we are working on contain 80 copies. This means that achieving homoplasmy (making all copies of the chloroplast genome the same, with the gene of interest inserted) can take months and require several rounds of selection. The additional complication and time required to transform algal chloroplasts makes this field less attractive for researchers, despite its vast potential. We aspired to create a toolkit containing genetic constructs and hardware that would enable the user to insert a gene of their choice into the algal chloroplast genome faster and more efficiently than is currently possible.
We considered and tried to streamline every step of the chloroplast engineering process – cloning, transformation, incubation and waiting for growth.
Step 1: clone
When we started our project, there were no parts in the iGEM registry designed for chloroplasts – in contrast to the 20,000 biobricks available for engineering other organisms. We designed a library of 18 parts following the phytobrick syntax, containing all the basic elements of genetic circuits – promoters, terminators, homology regions and reporters.
One of these parts was cas9 codon-optimised for Chlamydomonas. We designed a strategy using CRISPR-cas9 to cleave the chloroplast genome at specific sites, allowing for targeted insertion of our gene of interest using homologous recombination. This would theoretically enable homoplasmy to be achieved in one generation rather than taking several months.
Step 2: transform
‘Gene guns’ are one of the fastest, most efficient techniques out there for chloroplast transformation. A gene gun basically does what it says on the tin – fires metal particles coated with DNA into cells to genetically transform them, using bursts of high-pressure gas. The drawback of this technique is that gene guns cost around £25,000. Inspired by the DIY science community that we interacted with at the Biodesign Nightscience conference in Paris – and in keeping with our aim to increase the accessibility of chloroplast transformation – we built an open-source gene gun for just 1% of the cost. We have been in contact with community labs across the world – from Los Angeles to London to Berlin – who are keen to use our design.
Step 3: incubate
One of the problems we faced in the project was creating the right growth conditions to grow certain Chlamydomonas strains, so we built a growth facility to make this alga a more accessible chassis for future teams. It is fully customisable with light control, temperature regulation and imaging capabilities – the user can even monitor the growth of their algae from a distance as it automatically posts photos at regular intervals to its own Twitter account!
Step 4: wait
Using current techniques, it takes 2-3 months to achieve homoplasmy. We used kinetic modelling to illustrate how using our cas9 strategy would massively accelerate this process – achieving homoplasmy in just seven hours.
After four months of living and breathing this project, we found ourselves in Boston preparing for the iGEM Jamboree. It was difficult to appreciate the scale of what we had been involved in until we were sitting in a packed auditorium with thousands of other people from teams across the world who’d been doing the same thing. We were all inspired by the huge potential of synthetic biology shown in the variety of projects presented, and by what can be achieved in one summer if groups of enthusiastic, motivated people from different disciplines are brought together.
Then, very suddenly, it was all over. Jetlagged and facing the cold, hard reality of returning to a Cambridge term in full swing, we could have regretted the time and effort that has gone into this project. However, we have had the opportunity to essentially run our own lab and plan a project entirely from scratch – a level of independence that is rare for a group of undergraduates. We met a huge variety of people working in different disciplines, spread our ideas about our project and synthetic biology as a whole through articles and outreach events, and even had the opportunity to travel to Paris and Boston. We were rewarded for these efforts with a gold medal and the Best Plant Synthetic Biology prize at the Jamboree. During this event, we were even approached by a former iGEM judge who was interested to helping to distribute our gene gun to future iGEM teams in South America, and asked if we would be interested in travelling there next summer to mentor some of these teams. Therefore, although our wiki is complete, our presentation has been given and our poster stored away, the opportunities that iGEM has given us will continue to have an influence well beyond this summer.
Note from Cambridge Consultants
Synthetic biology has huge potential to solve many of today’s critical challenges in healthcare, agriculture, energy and the environment. That’s why Cambridge Consultants has decided to sponsor the Cambridge University team at iGEM 2016 – the international genetically engineered machine competition run by MIT. As part of our sponsorship, we are acting as mentors – giving the team access to more than 700 Cambridge Consultants engineers and scientists worldwide to help solve problems during this year’s project.
The iGEM team is also grateful for support from:
- University of Cambridge, School of Biological Sciences