Difference between revisions of "Part:BBa K4390002"

 
 
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<partinfo>BBa_K4390002 short</partinfo>
 
<partinfo>BBa_K4390002 short</partinfo>
  
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This construct assembled using JUMP is a working transcriptional unit for the transcriptional repressor ArsR. This compound part is used to transform cells to express ArsR, so that the cell lysate contains ArsR and can be used in our biosensor.
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BBa_K4390002 is a working transcriptional unit which expresses the ArsR repressor (BBa_J15101). It is composed of the constitutive promoter J23100  (BBa_J23100), a ribosome binding site (BBa_B0034), the arsR repressor sequence (BBa_J15101) and the weak synthetic L2U2H09 Terminator (BBa_K4390001). This unit was assembled using JUMP and can be assembled into a pJUMP29-1A KanR Type IIS Level 1 vector plasmid for expression in cells.  
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===Usage and Biology===
 
===Usage and Biology===
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The arsR repressor is derived from an Escherichia coli chromosomal arsR coding sequence which binds to the ars promoter and represses it in the absence of arsenate or arsenite. We used this transcriptional unit alongside the As biosensor (BBa_K3380600) so that when we use cell lysate which expresses our transcriptional unit there would be arsR present in the lysate. When this cell-free extract is combined with the As biosensor it induces transcriptional repression of the linear biosensor when arsenate or arsenite is not present. When arsenite or arsenate is present the arsR bind to the metal ions and allows transcription of the biosensor and would therefore generate a fluorescent output. It should be noted that T7 RNA polymerase, chemical energy (ATP), NTPs and the DFHBI are also required in the cell-free reaction so that fluorescence is observed.
  
 
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Latest revision as of 15:54, 8 September 2022

ArsR expression construct

BBa_K4390002 is a working transcriptional unit which expresses the ArsR repressor (BBa_J15101). It is composed of the constitutive promoter J23100 (BBa_J23100), a ribosome binding site (BBa_B0034), the arsR repressor sequence (BBa_J15101) and the weak synthetic L2U2H09 Terminator (BBa_K4390001). This unit was assembled using JUMP and can be assembled into a pJUMP29-1A KanR Type IIS Level 1 vector plasmid for expression in cells.


Usage and Biology

The arsR repressor is derived from an Escherichia coli chromosomal arsR coding sequence which binds to the ars promoter and represses it in the absence of arsenate or arsenite. We used this transcriptional unit alongside the As biosensor (BBa_K3380600) so that when we use cell lysate which expresses our transcriptional unit there would be arsR present in the lysate. When this cell-free extract is combined with the As biosensor it induces transcriptional repression of the linear biosensor when arsenate or arsenite is not present. When arsenite or arsenate is present the arsR bind to the metal ions and allows transcription of the biosensor and would therefore generate a fluorescent output. It should be noted that T7 RNA polymerase, chemical energy (ATP), NTPs and the DFHBI are also required in the cell-free reaction so that fluorescence is observed.

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 11
    Illegal NheI site found at 34
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 196
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]