Difference between revisions of "Part:BBa K2442101:Design"
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In absence of L-arabinose the AraC dimer binds to pBAD operator half-sites O<sub>2</sub> and I<sub>1</sub> and represses transcription by excluding RNA polymerase from binding to pBAD or PC. Binding of L-arabinose causes a conformational change in the protein such that the DNA-binding domains of the dimer bind to adjacent I<sub>1</sub and I<sub>2</sub> half-sites, resulting in transcription activation of downstream genes (Fig. 1).
 
In absence of L-arabinose the AraC dimer binds to pBAD operator half-sites O<sub>2</sub> and I<sub>1</sub> and represses transcription by excluding RNA polymerase from binding to pBAD or PC. Binding of L-arabinose causes a conformational change in the protein such that the DNA-binding domains of the dimer bind to adjacent I<sub>1</sub and I<sub>2</sub> half-sites, resulting in transcription activation of downstream genes (Fig. 1).
  
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[[Image:T-Glasgow-K2442101-design-Fig1.png|left|thumb|800px|'''Figure 1: Regulation of the L-arabinose operon by arabinose. In the absence of L-arabinose, AraC dimer binds to O<sub>2</sub> and I<sub>1</sub> half sites causing DNA looping, which prevents RNA polymerase from accessing P<sub>BAD</sub>. Upon binding of L–arabinose, the AraC dimer binds I<sub>1</sub> and I<sub>2</sub> half sites instead, allowing transcription of the polycistronic ''araBAD'' mRNA. ''araC'' is transcribed in opposite direction from ''araBAD'', and is under control of the PC promoter. RNA polymerase and AraC compete for binding at O<sub>1</sub> and P<sub>C</sub>. Adapted from Shleif (2000) <ref> Schleif, R. (2000). Regulation of the L-arabinose operon of ''Escherichia coli''. Trends In Genetics ''16'', 559-565..</ref>''']]
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[[Image:T-Glasgow-K2442101-design-Fig1.png|centre|thumb|800px|'''Figure 1: Regulation of the L-arabinose operon by arabinose. In the absence of L-arabinose, AraC dimer binds to O<sub>2</sub> and I<sub>1</sub> half sites causing DNA looping, which prevents RNA polymerase from accessing P<sub>BAD</sub>. Upon binding of L–arabinose, the AraC dimer binds I<sub>1</sub> and I<sub>2</sub> half sites instead, allowing transcription of the polycistronic ''araBAD'' mRNA. ''araC'' is transcribed in opposite direction from ''araBAD'', and is under control of the PC promoter. RNA polymerase and AraC compete for binding at O<sub>1</sub> and P<sub>C</sub>. Adapted from Shleif (2000) <ref> Schleif, R. (2000). Regulation of the L-arabinose operon of ''Escherichia coli''. Trends In Genetics ''16'', 559-565..</ref>''']]
  
  

Revision as of 23:25, 1 November 2017


Minimal pBAD promoter


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BamHI site found at 241
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal AgeI site found at 76
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal SapI site found at 58


Design Notes

In absence of L-arabinose the AraC dimer binds to pBAD operator half-sites O2 and I1 and represses transcription by excluding RNA polymerase from binding to pBAD or PC. Binding of L-arabinose causes a conformational change in the protein such that the DNA-binding domains of the dimer bind to adjacent I1</sub and I2 half-sites, resulting in transcription activation of downstream genes (Fig. 1).

Figure 1: Regulation of the L-arabinose operon by arabinose. In the absence of L-arabinose, AraC dimer binds to O2 and I1 half sites causing DNA looping, which prevents RNA polymerase from accessing PBAD. Upon binding of L–arabinose, the AraC dimer binds I1 and I2 half sites instead, allowing transcription of the polycistronic araBAD mRNA. araC is transcribed in opposite direction from araBAD, and is under control of the PC promoter. RNA polymerase and AraC compete for binding at O1 and PC. Adapted from Shleif (2000) [1]


Source

PCR amplification from BBa_I0500.

===References===

  1. ↑ Schleif, R. (2000). Regulation of the L-arabinose operon of Escherichia coli. Trends In Genetics 16, 559-565..