Difference between revisions of "Team:Oxford/Synbiota"

We were awarded trailblazer scholarship by Synbiota, a startup company co-founded by iGEM alumni, and had a chance to test their prototype Rapid DNA Prototyping (RDP) assembly standard including exclusive iGEM kit and RDP part designing software.

RDP is a modular approach to synthetic biology that is biobrick compatible, and somewhat similar to standard biobrick approach in a sense that DNA templates can create new RDP parts through PCR. However, each RDP part is defined by each promoter, RBS, coding region, or terminator. Series of RDP parts can be assembled using magnetic anchor under two hours.

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RDP Standard requires addition of two unique sticky ends (X and Z) and their complements (X’ and Z’) onto each part, allowing sequential assembly from promoter to terminator.

We have designed two customised RDP parts, pLsr and T4Holin. These RDP parts were designed using GEntle software to contain X-Z’ or Z-X’ sticky ends. Synbiota has provided us with physical RDP parts of pLse and T4Holin, as well as primers to amplify them through PCR. Wetlab notebooks of Synbiota specific PCR can be found here

Then pLsr and T4Holin were combined with provided Anchor RDP part, RBS, terminator and Cap to make a full RDP circuit which is equivalent to BBa_K1659601.

So this is how our part BBa_K1659601 RDP circuit looks like:

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Each ends of Anchor and Cap RDP parts self-assemble to form pSB1C3 backbone that is biobrick compatible.

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RDP assembly method is very time-efficient because the assembly starts with a Synbiota Anchor part (DNA anchored to a magnetic bead), and first RDP part (pLsr in our case) is added onto the solution, requiring ligation incubation time of only 8min. Then, an eppendorf tube holder with the magnet pellets Anchor-pLsr component, washing off excess DNA and ligase. The picture below illustrates this very stage.

References

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