Difference between revisions of "Part:BBa K2918032"

 
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<partinfo>BBa_K2918032 short</partinfo>
 
<partinfo>BBa_K2918032 short</partinfo>
  
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A level 1 with 0.5 T7 promoter, a universal RBS, a GFP and a T7 terminator
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A level 1 with Medium T7 promoter, a universal RBS, a GFP and a T7 terminator
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===Usage and Biology===
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<span class='h3bb'>Sequence and Features</span>
 
<span class='h3bb'>Sequence and Features</span>
 
<partinfo>BBa_K2918032 SequenceAndFeatures</partinfo>
 
<partinfo>BBa_K2918032 SequenceAndFeatures</partinfo>
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The construct is confirmed by sequencing and there are no mutations.
  
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===Toxicity===
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Our Sci-Phi 29 tool is based on four components of the Φ29 bacteriophage: DNAP, TP, p5 and p6. However, overexpression of these proteins are toxic for the cell. In order to determine the optimal expression levels of the proteins in live cells, we carried out viability assays in <i>E. coli</i> BL21(DE3) pLysS. The results are shown in the graphs below.
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<div><ul>
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  <li style="display: inline-block;"> [[File:T--TUDelft--0.5gfp-nomM.png|thumb|none|444px|<b>Figure 3A:</b> The growth curve of GFP under a medium T7 promoter in hours with no IPTG induction]] </li>
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  <li style="display: inline-block;"> [[File:T--TUDelft--0.5gfp-1mM.png|thumb|none|444px|<b>Figure 3B:</b> The growth curve of GFP under a medium T7 promoter in hours with 1 mM IPTG induction]] </li>
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    <li> [[File:T--TUDelft--0.5gfp-10mM.png|thumb|center|444px|<b>Figure 3C:</b> The growth curve of GFP under a medium T7 promoter in hours with 10 mM IPTG inductionn]] </li>
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    </ul></div>
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<!-- Add more about the biology of this part here
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===Usage and Biology===
  
 
<!-- Uncomment this to enable Functional Parameter display  
 
<!-- Uncomment this to enable Functional Parameter display  

Latest revision as of 22:25, 6 December 2019


Medium T7 promoter - Universal RBS - GFP - T7 terminator

A level 1 with Medium T7 promoter, a universal RBS, a GFP and a T7 terminator

Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal EcoRI site found at 465
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 465
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 465
    Illegal BamHI site found at 674
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal EcoRI site found at 465
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal EcoRI site found at 465
  • 1000
    COMPATIBLE WITH RFC[1000]

The construct is confirmed by sequencing and there are no mutations.

Toxicity

Our Sci-Phi 29 tool is based on four components of the Φ29 bacteriophage: DNAP, TP, p5 and p6. However, overexpression of these proteins are toxic for the cell. In order to determine the optimal expression levels of the proteins in live cells, we carried out viability assays in E. coli BL21(DE3) pLysS. The results are shown in the graphs below.

  • Figure 3A: The growth curve of GFP under a medium T7 promoter in hours with no IPTG induction
  • Figure 3B: The growth curve of GFP under a medium T7 promoter in hours with 1 mM IPTG induction
  • Figure 3C: The growth curve of GFP under a medium T7 promoter in hours with 10 mM IPTG inductionn