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Team:Bielefeld-Germany/Results/S-Layer/Guide/2
From 2011.igem.org
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[[Image:IGEM-Bielefeld2011-Wirbeimklonen.JPG|300px|thumb|left|iGEM assembly line]] | [[Image:IGEM-Bielefeld2011-Wirbeimklonen.JPG|300px|thumb|left|iGEM assembly line]] | ||
[[Image:IGEM-Bielefeld2011-Vieleplatten.JPG|400px|thumb|left|Clone wars]] | [[Image:IGEM-Bielefeld2011-Vieleplatten.JPG|400px|thumb|left|Clone wars]] | ||
| + | [[Image:Bielefeld-Germany-2011-Simple-cloning-example.png|400px|thumb|Freiburg Assembly example]] | ||
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To create fusion proteins you have to use a BioBrick assembly which assembles in frame so your fusion protein is functional. The standard BioBrick assembly '''does not assemble in frame''' so you have to use other standards - like the Freiburg or the Gibson assembly. Our S-layer gene is already provided in the [http://partsregistry.org/Assembly_standard_25 Freiburg assembly standard] so after ordering it you can directly start assembling your fusion proteins of choice to it. In addition, we provided these genes not only in this easy to use standard, but they also exist under the control of a T7 / lac promoter (<partinfo>K525303</partinfo> and <partinfo>K525403</partinfo>). So after cloning you can easily express them in an ''E. coli'' strain which carries a T7 polymerase in its genome like BL21(DE3) or KRX. Do not forget to add a '''his-tag''' (<partinfo>K157011</partinfo>) to your fusion protein so you can easily purify it and follow the next steps in this guide! | To create fusion proteins you have to use a BioBrick assembly which assembles in frame so your fusion protein is functional. The standard BioBrick assembly '''does not assemble in frame''' so you have to use other standards - like the Freiburg or the Gibson assembly. Our S-layer gene is already provided in the [http://partsregistry.org/Assembly_standard_25 Freiburg assembly standard] so after ordering it you can directly start assembling your fusion proteins of choice to it. In addition, we provided these genes not only in this easy to use standard, but they also exist under the control of a T7 / lac promoter (<partinfo>K525303</partinfo> and <partinfo>K525403</partinfo>). So after cloning you can easily express them in an ''E. coli'' strain which carries a T7 polymerase in its genome like BL21(DE3) or KRX. Do not forget to add a '''his-tag''' (<partinfo>K157011</partinfo>) to your fusion protein so you can easily purify it and follow the next steps in this guide! | ||
Revision as of 01:02, 29 October 2011
Assembly of functionalized S-layer proteins
To create fusion proteins you have to use a BioBrick assembly which assembles in frame so your fusion protein is functional. The standard BioBrick assembly does not assemble in frame so you have to use other standards - like the Freiburg or the Gibson assembly. Our S-layer gene is already provided in the [http://partsregistry.org/Assembly_standard_25 Freiburg assembly standard] so after ordering it you can directly start assembling your fusion proteins of choice to it. In addition, we provided these genes not only in this easy to use standard, but they also exist under the control of a T7 / lac promoter (<partinfo>K525303</partinfo> and <partinfo>K525403</partinfo>). So after cloning you can easily express them in an E. coli strain which carries a T7 polymerase in its genome like BL21(DE3) or KRX. Do not forget to add a his-tag (<partinfo>K157011</partinfo>) to your fusion protein so you can easily purify it and follow the next steps in this guide!
Want to know how to express S-layer fusion proteins based on <partinfo>K525303</partinfo> the simplest way? Then click here.
Or do you want to produce bigger amounts of your S-layer to coat all the things? Then get the big one here.
Have problems with your assemblies? Check out our genetic engineering protocols.
