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Team:TU Munich/Project/Vector Design
From 2012.igem.org
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| + | vom Poster: | ||
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| + | RFC25 COMPATIBILITY | ||
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| + | To be able to test and quantify the expression of desired enzymes in yeast we designed an expression vector which is compatible to the iGEM RFC25 standard based on the commercially available pYES2 vector from Invitrogen. | ||
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| + | DESIGN | ||
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| + | This included an exchange of the original multiple cloning site by a multiple cloning site consisting of the restriction enzymes used in the iGEM RFC25 standard. Additionally the restriction sites of these enzymes in the vector backbone have been eliminated by quick-change mutagenisis. Furthermore we inserted a gene sequence coding for a strep-tag II straight behind the multiple cloning site in order to facilitate the extraction of expressed proteins. The successful introduction of these modifications has been confirmed via DNA sequencing. | ||
| + | Additionally a second vector containing a His-tag instead of the strep-tag II and a vector giving the opportunity to test different promoters is currently planned. | ||
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Description of the original vector + 4 different modified versions. | Description of the original vector + 4 different modified versions. | ||
</div> | </div> | ||
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== Characterization == | == Characterization == | ||
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Revision as of 19:42, 24 September 2012
Contents |
Vector Design
Motivating text why we choose pYES2 and why we wanted to introduce a new vector to the registry.
Design
vom Poster:
RFC25 COMPATIBILITY
To be able to test and quantify the expression of desired enzymes in yeast we designed an expression vector which is compatible to the iGEM RFC25 standard based on the commercially available pYES2 vector from Invitrogen.
DESIGN
This included an exchange of the original multiple cloning site by a multiple cloning site consisting of the restriction enzymes used in the iGEM RFC25 standard. Additionally the restriction sites of these enzymes in the vector backbone have been eliminated by quick-change mutagenisis. Furthermore we inserted a gene sequence coding for a strep-tag II straight behind the multiple cloning site in order to facilitate the extraction of expressed proteins. The successful introduction of these modifications has been confirmed via DNA sequencing. Additionally a second vector containing a His-tag instead of the strep-tag II and a vector giving the opportunity to test different promoters is currently planned.
Description of the original vector + 4 different modified versions.
