About This Project
Millions of people die from viral infections every year. To work towards a cure we are studying the fundamental mechanisms of the infectious process. This project will define the minimal set of viral proteins required for starting its life cycle. We will engineer the genome of both the vaccinia virus and a complementing host strain to define this essential gene set. This work will lead to the development of entirely new biotechnologies and health applications.
Ask the Scientists
Join The DiscussionWhat is the context of this research?
Several studies have characterised the total proteome of purified vaccinia virus. These studies list all of the proteins that the virus is made from. Computational analysis has identified homologies and predicted functions for several of these proteins. Currently, we do not have functional proof of the specific proteins that are essential for the initiation of viral replication. This knowledge will give us a more complete picture of the early transcriptional and translational processes. We will then be able to understand how the viral machinery allocates its limited resources to simultaneously propagate and evade the host defence mechanisms when it is most vulnerable.
A similar experimental approach has been taken before for single genes but never to reveal complete gene sets.
What is the significance of this project?
A complete understanding of the molecular mechanisms and proteins involved in the early stages of viral infection is key to understanding the complex host-pathogen interactions that lead to disease progression. Our functional inventory will advance the future development therapeutic interventions.
In addition to advancing academic knowledge, we believe that our work will also be useful for viral engineers. Currently, vaccinia is being used to target cancer and as a delivery vector for the next generation of vaccines.
Vaccinia is used for the recombinant production of proteins in mammalian cells. This work will directly lead to an improved expression system where eventually we will be able to completely circumvent the use of the live virus.
What are the goals of the project?
We will use our new synthetic genomic technologies to manipulate the genomes of both vaccinia and the host strain to isolate a functional gene set. Taking a systematic gene deletion/complementation approach followed by a phenotypic characterisation, we will define all of the essential genes within vaccinia that are required to kickstart the infectious lifecycle.
We have already begun some of the computational and experimental designs for this project.
I will directly supervise and work with the engineer that is hired for this project. We will both be working full-time cloning, culturing cells, propagating virus, refactoring the expression of entire gene sets and constructing stable cell lines. We will additionally use microscopy and qPCR for characterisation.
Budget
The cost will be for the salary of an experienced full-time engineer that we will need to help us with the extensive cell culture and molecular biology work. The research contract will be for one year. This project will be extremely labor intensive. Once we have the cell cultures going we have to work everyday, including weekends, to maintain them. It will take two workers to cover the demanding schedule. The funds requested will cover the compulsory French employer paid social contribution which is 42% of the brut salary.
Additionally, we expect that this project will use a considerable amount of consumables because it entails a heavy load of molecular biology, sequencing, cell culture and synthetic DNA assembly.
The cost for consumables that will be used in this project will be paid for by us using resources that we currently have allocated for this in our budgets. We estimate that this expense will be in the range of 15,000-20,000€ for a one year project.
Endorsed by
Meet the Team
Brian Jester
The engineering and re-design of life has fascinated me for years. In this exciting project, we now have the opportunity to systematically tease out the players involved with the early stages of viral infection.
I have undertaken a diverse professional course which includes both formal education and work experiences that span several disciplines within industry and academia.
I obtained my undergraduate and Masters degrees in Biology from University of Wisconsin. I then secured a scholarship to do my PhD in Genetics at Trinity College, Dublin Ireland (awarded 2006). I have 10 publications, 1 research grant funded and contributed to 3 patents. During my PhD, my fascination with genome engineering really took off. Quickly, I came to learn that DNA is just the text of the blueprint for life that is entirely re-writable by us, the engineers.
The learning path has been extremely diverse. I've held several Postdoctoral positions (10 years) in three different disciplines, worked in Industrial research and have started up two successful companies. It was during my second and third postdocs that I started working with vaccinia. Our work resulted in an improved method for the recombinant production of proteins in mammalian cells.
The last 5 years I have focussed my energies on the field of Synthetic Biology. We have made significant advances in our understanding of how gene layout influences expression. We have invented new molecular methods and tools for genome engineering. All of the resources and expertise that we have developed over these long years will be put forward into ensuring that this project will be a success.
Lab Notes
Nothing posted yet.
Additional Information
Experimentally: Our work builds on the published fowlpox rescue system that has been used to "kick start" a viral genome from a BAC. By engineering a host strain that can independently complement the vBAC without the need of a helper virus will be our goal.
We will begin by cloning the entire vaccinia genome into a vector. The introduction of a constitutively expressed GFP gene within the vaccinia genome will allow us to monitor viral growth in vivo. We will then insert the viral genes that are predicted to be involved with transcription into the host strain chromosome using the piggyBAC system. To evaluate the rescue of the virus from the transfected vBAC we will use fluorescent microscopy to visualise viral propagation.
This tool will prove to be a spring board platform that will allow us to determine functionality of other gene sets using a similar deletion/complementation approach.
Project Backers
- 5Backers
- 2%Funded
- $815Total Donations
- $163.00Average Donation