Difference between revisions of "Part:BBa K2918020"

 
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A level 1 construct with a 0.1 T7 p, a universal RBS, DNAP and a T7 terminator.
 
A level 1 construct with a 0.1 T7 p, a universal RBS, DNAP and a T7 terminator.
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===Usage and Biology===
 
  
 
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<partinfo>BBa_K2918020 SequenceAndFeatures</partinfo>
 
<partinfo>BBa_K2918020 SequenceAndFeatures</partinfo>
  
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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--0.1DNAP-no.png|thumb|none|444px|<b>Figure 3A:</b> The growth curve of phi 29 DNAP with a 0.1 T7 promoter in hours with no IPTG induction]] </li>
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    <li style="display: inline-block;"> [[File:T--TUDelft--0.1DNAP-0.001mM.png|thumb|none|444px|<b>Figure 3B:</b> The growth curve of phi 29 DNAP with a 0.1 T7 promoter in hours with 0.001 mM IPTG induction]] </li>
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    <li style="display: inline-block;"> [[File:T--TUDelft--0.1DNAP-0.01mM.png|thumb|none|444px|<b>Figure 3C:</b> The growth curve of phi 29 DNAP with a 0.1 T7 promoter in hours with 0.01 mM IPTG induction]] </li>
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    <li style="display: inline-block;"> [[File:T--TUDelft--0.1DNAP-0.1mM.png|thumb|none|444px|<b>Figure 3D:</b> The growth curve of phi 29 DNAP with a 0.1 T7 promoter in hours with 0.1 mM IPTG induction]] </li>
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    <li style="display: inline-block;"> [[File:T--TUDelft--0.1DNAP-1mM.png|thumb|none|444px|<b>Figure 3E:</b> The growth curve of phi 29 DNAP with a 0.1 T7 promoter in hours with 1 mM IPTG induction]] </li>
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    <li style="display: inline-block;"> [[File:T--TUDelft--0.1DNAP-10mM.png|thumb|none|444px|<b>Figure 3F:</b> The growth curve of phi 29 DNAP with a 0.1 T7 promoter in hours with 10 mM IPTG induction]] </li>
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    </ul></div>
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===Usage and Biology===
  
 
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Revision as of 08:29, 4 October 2019


Weak T7 promoter - Universal RBS - Φ29 DNAP - T7 terminator

A level 1 construct with a 0.1 T7 p, a universal RBS, DNAP and a T7 terminator.

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal AgeI site found at 1709
  • 1000
    COMPATIBLE WITH RFC[1000]

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 phi 29 DNAP with a 0.1 T7 promoter in hours with no IPTG induction
  • Figure 3B: The growth curve of phi 29 DNAP with a 0.1 T7 promoter in hours with 0.001 mM IPTG induction
  • Figure 3C: The growth curve of phi 29 DNAP with a 0.1 T7 promoter in hours with 0.01 mM IPTG induction
  • Figure 3D: The growth curve of phi 29 DNAP with a 0.1 T7 promoter in hours with 0.1 mM IPTG induction
  • Figure 3E: The growth curve of phi 29 DNAP with a 0.1 T7 promoter in hours with 1 mM IPTG induction
  • Figure 3F: The growth curve of phi 29 DNAP with a 0.1 T7 promoter in hours with 10 mM IPTG induction