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Team:Washington/Project/CDS
From 2009.igem.org
(New page: __NOTOC__ {{Template:Team:Washington/Templates/Header}} ===Custom Display Vector=== ====Problem==== The underlying design of the streptavidin display constructs in the registry has the d...) |
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The underlying design of the streptavidin display constructs in the registry has the displayed protein bound close to the cell membrane. This constraint could be preventing streptavidin on the surface of the cell from forming tetramers lowering its effectiveness at binding biotin. Furthermore these existing display systems prevent the addition of another protein into them, preventing the user from displaying other proteins. | The underlying design of the streptavidin display constructs in the registry has the displayed protein bound close to the cell membrane. This constraint could be preventing streptavidin on the surface of the cell from forming tetramers lowering its effectiveness at binding biotin. Furthermore these existing display systems prevent the addition of another protein into them, preventing the user from displaying other proteins. | ||
Revision as of 03:30, 19 October 2009
Custom Display System
Problem
The underlying design of the streptavidin display constructs in the registry has the displayed protein bound close to the cell membrane. This constraint could be preventing streptavidin on the surface of the cell from forming tetramers lowering its effectiveness at binding biotin. Furthermore these existing display systems prevent the addition of another protein into them, preventing the user from displaying other proteins.
Idea
The first goal of making a new display vector was to incorporate a GS linker between the displayed protein and the ompA protein anchoring in in the cell wall. However when reviewing the construct we decided to add some other useful features as well. Maintaining the Lpp tag (to direct the protein to the periplasm) and the OmpA trans-membrane regions (for anchoring the construct) of the original 2006 parts we added:
- GS Linker (Gly4Ser)4 - allowing for more space between the protein and the cell wall
- TEV (Tyrosine-Glutamate-Valine) - protease site allowing for cleavage of displayed proteins
- NheI restriction site - allowing for the insert of any BioBrick protein into the display construct
Current Status
We have built the Custom Display Vector and have inserted streptavidin into it, however we ran out of time and were not able to characterize this part. It was subbmited to the registry with streptavidin (BBa_K215210 and BBa_K215211) and without a displayed protein(BBa_K215200 and BBa_K215200).
