Difference between revisions of "Team:Lethbridge/Improve"

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<p><b>designed by:</b> Bartholomew Canton</p>
 
<p><b>designed by:</b> Bartholomew Canton</p>
 
<p><b>Rational behind improvements:</b></p>
 
<p><b>Rational behind improvements:</b></p>
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<p>Encodes only for the coding region of T7 RNA polymerase. In order to improve it and incorporate it into our Next<i>Vivo</i> system we have codon optimized it and attached a 6 X His tag for easy purification. </P>   
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<p>BBa_I2032 encodes exclusively for the coding region of T7 RNA polymerase. In order to improve it and incorporate it into our Next<i>Vivo</i> system we have Codon optimized for use in <i>Escherichia coli</i> and attached a N-terminal 6 X His tag with a serine glycine linker for easy purification. We have also optimized our T7 RNA polymerase to be overexpressed in BL21DE3 gold cells by putting it under the control of a T7 promoter (BBa_I719005), RBS (BBa_B0034) and double terminator (BBa_B0015)</P>   
  
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                    <h2>Part BBa_K1791002</h2>
 
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<p>Part BBa_K17910012 contains two modules, a low efficiency ribosome binding site (RBS)  expressing MS2 coat protein and a theophylline ribozyme with a MS2 aptamer at the 3’ end. To express  MS2 coat protein the part utilizes expression of c-terminal 6x His tagged MS2 coat protein under a T7  promoter and a standard Shine Dalgarno sequence followed by a double terminator (BBa_0015).  Downstream of the RBS is a T7 promoter followed by a theophylline ribozyme and a MS2 aptamer at the  3’ end. Coexpression of the two modules results in MS2 binding to the 3’ end of the theophylline  ribozyme construct which allows for MS2-RNA purification by nickel affinity chromatography, RNA can  then be eluted from the MS2-RNA complex by addition of theophylline.</p>
 
  
 
                 </div>
 
                 </div>

Revision as of 05:55, 19 October 2017

Improved Parts

BBa_K2443037 - T7 RNA Polymerase

Original part: BBa_I2032

Submitted by: MIT in 2006

designed by: Bartholomew Canton

Rational behind improvements:

BBa_I2032 encodes exclusively for the coding region of T7 RNA polymerase. In order to improve it and incorporate it into our NextVivo system we have Codon optimized for use in Escherichia coli and attached a N-terminal 6 X His tag with a serine glycine linker for easy purification. We have also optimized our T7 RNA polymerase to be overexpressed in BL21DE3 gold cells by putting it under the control of a T7 promoter (BBa_I719005), RBS (BBa_B0034) and double terminator (BBa_B0015)