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Team:DTU Denmark/USERprinciple
From 2009.igem.org
The USER fusion assembly standard allows rapid construction of multi-part devices, without some of the drawbacks of the restriction-enzyme based standard biobrick assembly method. The main advantages of this assembly method is:
1. Standardized method of linking multiple biobricks together.
2. Nearly all restriction sites are allowed in the biobricks.
3. 8 basepair-overhangs allows ligase-free cloning. With this protocol E. coli can be transformed with a multipart-construct less than 2 hours after your PCR-reaction has completed.
4. USER fusion allows for scarfree fusions of DNA which is ideal for fusing protein domain biobricks.
5. Insertions of small sequences between biobricks such as a nucleous localization signal or Gly-Ser-Gly (GSG) flexible linker is possible with the right primer design.
5. High fidelity is ensured by using PfuTurbo® Cx Hotstart DNA polymerase.
6. By the design of the PCR tails, it can be decided whether the USER cassette should be deleted, copied or moved following insertion.
7. Directionallity of inserts are supported
