Difference between revisions of "Part:BBa I759005"

 
 
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As the recombineering, testing of riboregulators, and titering processes took place concurrently, we needed to find a simpler way to regulate viral protein concentrations in the cells. To this end, E. coli strains have been constructed that contain a low-copy plasmid construct where one of three key developmental viral genes - coding for the cro, N, or Q proteins - is regulated by a tetracycline-dependent promoter. A constitutive promoter produces a steady stream of tetracycline repressor (tetR), which substitutes for the cis repressor in repressing protein levels. The addition of anhydrotetracycline (aTc, acting as the trans activator) inactivates the tetracycline repressor and leads to the production of the respective viral protein in the E. coli cells. This allows us to control the concentration of viral protein produced in the cells by adding varying amounts of aTc to the bacterial growth media.
  
 
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===Functional Parameters===
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==Functional Parameters: Austin_UTexas==
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<partinfo>BBa_I759005 parameters</partinfo>
 
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<h3><center>Burden Imposed by this Part:</center></h3>
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<center><img src = "https://static.igem.org/mediawiki/parts/f/fa/T--Austin_Utexas--no_burden_icon.png" style = "width:160px;height:120px"></center>
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<figcaption><center><b>Burden Value: 17.8 ± 16% </b></center></figcaption>
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<p> Burden is the percent reduction in the growth rate of <i>E. coli</i> cells transformed with a plasmid containing this BioBrick (± values are 95% confidence limits). This BioBrick did not exhibit a burden that was significantly greater than zero (i.e., it appears to have little to no impact on growth). Therefore, users can depend on this part to remain stable for many bacterial cell divisions and in large culture volumes. Refer to any one of the
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<a href="https://parts.igem.org/Part:BBa_K3174002">BBa_K3174002</a> - <a href="https://parts.igem.org/Part:BBa_K3174007">BBa_K3174007</a> pages for more information on the methods, an explanation of the sources of burden,  and other conclusions from a large-scale measurement project conducted by the <a href="http://2019.igem.org/Team:Austin_UTexas">2019 Austin_UTexas team</a>.</p>
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<p>This functional parameter was added by the <a href="https://2020.igem.org/Team:Austin_UTexas/Contribution">2020 Austin_UTexas team.</a></p>
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Latest revision as of 22:05, 27 August 2020

tet-regulated cro (Weak tetR promoter)

As the recombineering, testing of riboregulators, and titering processes took place concurrently, we needed to find a simpler way to regulate viral protein concentrations in the cells. To this end, E. coli strains have been constructed that contain a low-copy plasmid construct where one of three key developmental viral genes - coding for the cro, N, or Q proteins - is regulated by a tetracycline-dependent promoter. A constitutive promoter produces a steady stream of tetracycline repressor (tetR), which substitutes for the cis repressor in repressing protein levels. The addition of anhydrotetracycline (aTc, acting as the trans activator) inactivates the tetracycline repressor and leads to the production of the respective viral protein in the E. coli cells. This allows us to control the concentration of viral protein produced in the cells by adding varying amounts of aTc to the bacterial growth media.

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 464
    Illegal NheI site found at 487
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 174
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]



Functional Parameters: Austin_UTexas

BBa_I759005 parameters

Burden Imposed by this Part:

Burden Value: 17.8 ± 16%

Burden is the percent reduction in the growth rate of E. coli cells transformed with a plasmid containing this BioBrick (± values are 95% confidence limits). This BioBrick did not exhibit a burden that was significantly greater than zero (i.e., it appears to have little to no impact on growth). Therefore, users can depend on this part to remain stable for many bacterial cell divisions and in large culture volumes. Refer to any one of the BBa_K3174002 - BBa_K3174007 pages for more information on the methods, an explanation of the sources of burden, and other conclusions from a large-scale measurement project conducted by the 2019 Austin_UTexas team.

This functional parameter was added by the 2020 Austin_UTexas team.