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Team:Potsdam Bioware/Project/Collaboration
From 2012.igem.org
Collaboration
with University of Freiburg, Germany
Introduction
As our team aims to mutate the antibody genes specifically and team Freiburg standardizes the TAL proteins that bind DNA specifically, we thought it would be a great idea to combine our efforts.
We are helping the University Freiburg team to test their Transactivator-like (TAL) proteins by sending one of our modified AID enzymes for ligation to the TAL domain. Team Freiburg, on the other hand, plans to send us back the TAL sequence ligated to the AID. Afterwards, we intend to check the mutation rate of the antibody sequence after directing the AID specifically (with help of TAL protein) to it.
A TAL domain is a DNA-binding protein which can bind specifically to a 14 base pair long target DNA sequence. In case of our project, this sequence is located close to the CDR3 region (framework) of the antibody fragment and enables binding of the TAL protein to our construct. Due to the conservation of this region in antibodies, the TAL protein created by team Freiburg should bind to common antibody sequences.
By linking the N-terminus of the AID to the TAL domain by a glycine-serine linker the enzyme AID can easily be brought in close proximity to the sequence that needs to be mutated. In this way we plan to obtain a more targeted directing of the enzyme and a higher mutation rate.
Having ligated the TAL protein sequence with AID sequence in one ORF, together we are creating AID protein that specifically binds to the antibody sequence and enables to mutate it.
Results of the directed AID mutation
We are waiting for Team Freiburg to send us the TAL domain linked to the AID to continue the test on it.
