Difference between revisions of "Part:BBa K4390097"
 
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This part is an improvement of [[Part:BBa_K3380600]], as it expands the possibility of usage to detecting mercury.  
 
This part is an improvement of [[Part:BBa_K3380600]], as it expands the possibility of usage to detecting mercury.  
  
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The Edinburgh-UHAS_Ghana team for 2022 designed a construct to detect lead in contaminated water. This construct is composed of a fluorescent RNA aptamer iSpinach flanked by the F30 tRNA scaffolds (BBa_K4390094) which under a strong T7 RNA promoter (BBa_I712074), downstream of the promotor there is the mercury promotor which acts as the binding site for the MerR repressor (BBa_K346002) as to allow transcriptional repression of the T7 promotor. This construct is designed to be cell-free and only requires transcription of the RNA aptamer to produce fluorescence. It should be noted that T7 RNA polymerase, chemical energy (ATP), NTPs and DFHBI or DFHO are also required in the cell-free reaction so that fluorescence is observed.
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The Edinburgh-UHAS_Ghana team for 2022 designed a construct to detect mercury in contaminated water. This construct is composed of a fluorescent RNA aptamer iSpinach flanked by the F30 tRNA scaffolds (BBa_K4390094) which under a strong T7 RNA promoter (BBa_I712074), downstream of the promotor there is the mercury promotor which acts as the binding site for the merR repressor (BBa_K346002) as to allow transcriptional repression of the T7 promotor. This construct is designed to be cell-free and only requires transcription of the RNA aptamer to produce fluorescence. It should be noted that T7 RNA polymerase, chemical energy (ATP), NTPs and DFHBI or DFHO are also required in the cell-free reaction so that fluorescence is observed.
  
  
 
===Usage and Biology===
 
===Usage and Biology===
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This biosensor should be used with a cell-free lysate which contains the MerR repressor, T7 RNA polymerase, chemical energy (ATP), NTPs and DFHBI or DFHO to induce fluorescence in the presence of mercury ions. The MerR repressor will bind to the MerR binding site on the mercury promoter when mercury ions are not present in the reaction. This causes transcriptional repression of the RNA aptamer as the T7 polymerase is physically occluded from reading the linear construct. When mercury ions are present in the reaction the pbrR repressor will then bind to the mercury ions and it will allow the transcription of the RNA aptamer, this aptamer can then bind to the DFHBI or DFHO which will induce fluorescence.  
+
This biosensor should be used with a cell-free lysate which contains the merR repressor, T7 RNA polymerase, chemical energy (ATP), NTPs and DFHBI or DFHO to induce fluorescence in the presence of mercury ions. The merR repressor will bind to the merR binding site on the mercury promoter when mercury ions are not present in the reaction. This causes transcriptional repression of the RNA aptamer as the T7 polymerase is physically occluded from reading the linear construct. When mercury ions are present in the reaction the pbrR repressor will then bind to the mercury ions and it will allow the transcription of the RNA aptamer, this aptamer can then bind to the DFHBI or DFHO which will induce fluorescence.  
 
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Latest revision as of 13:49, 12 October 2022


merR controlled iSpinach biosensor

Improvement of part BBa_K3380600

This part is an improvement of Part:BBa_K3380600, as it expands the possibility of usage to detecting mercury.

The Edinburgh-UHAS_Ghana team for 2022 designed a construct to detect mercury in contaminated water. This construct is composed of a fluorescent RNA aptamer iSpinach flanked by the F30 tRNA scaffolds (BBa_K4390094) which under a strong T7 RNA promoter (BBa_I712074), downstream of the promotor there is the mercury promotor which acts as the binding site for the merR repressor (BBa_K346002) as to allow transcriptional repression of the T7 promotor. This construct is designed to be cell-free and only requires transcription of the RNA aptamer to produce fluorescence. It should be noted that T7 RNA polymerase, chemical energy (ATP), NTPs and DFHBI or DFHO are also required in the cell-free reaction so that fluorescence is observed.


Usage and Biology

This biosensor should be used with a cell-free lysate which contains the merR repressor, T7 RNA polymerase, chemical energy (ATP), NTPs and DFHBI or DFHO to induce fluorescence in the presence of mercury ions. The merR repressor will bind to the merR binding site on the mercury promoter when mercury ions are not present in the reaction. This causes transcriptional repression of the RNA aptamer as the T7 polymerase is physically occluded from reading the linear construct. When mercury ions are present in the reaction the pbrR repressor will then bind to the mercury ions and it will allow the transcription of the RNA aptamer, this aptamer can then bind to the DFHBI or DFHO which will induce fluorescence. 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
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]