Part:BBa_K567013

tRNA(Asp)-TAG

tRNA(Asp) with its anticodon mutated to CUA (base pairing stop codon UAG) and under the control of lpp promoter. This biobrick is constructed first by cloning the tRNA(Asp) from AspV in E.coli, then the anticodon region is site-directed mutated.

Functional Parameters

chassis

Escherichia coli

Construction of BBa_K567013

This biobrick is constructed first by cloning the tRNA^Asp from AspV in E.coli, then the anticodon region is site-directed mutated. This part is constructed on the backbone plasmid pACYC184.

Characterization of BBa_K567013

This part acts as a stop codon suppressor tRNA. It can be charged with Asp.

We have used Pbla-Luc-TAG as our Reporter. The amount of luciferase produced is reflected using the bioluminescence emitted during the luciferin reaction. Our results demonstrate that this part can suppress TAG insertion into luciferase. In the experimental group, with the help of BBa_K567011 PT7-TDRS (Favorite Part), luciferase-TAG was produced and bioluminescence was emitted during the luciferin reaction. These results proved that this tRNA^Asp can effectively suppress stop codon.

fig. Functional Analysis of Stop-Codon Switch. ER2566 cannot produce luciferase with BBa_K567003 Pbla-Luc-TAG only. When BBa_K567011 PT7-TDRS (Favorite Part) and BBa_K567013 tRNA^Asp-TAG (Favorite Part) are also transformed into the cell, luciferase is produced. The results proved Stop-Codon Switch as a strict molecular switch without background noise.This tRNA^Asp is part of the Stop-Codon Switch

Sequence and Features

Assembly Compatibility:

• 10
  INCOMPATIBLE WITH RFC[10]

  Illegal EcoRI site found at 281
  
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal EcoRI site found at 281
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal EcoRI site found at 281
  
• 23
  INCOMPATIBLE WITH RFC[23]

  Illegal EcoRI site found at 281
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal EcoRI site found at 281
  
• 1000
  COMPATIBLE WITH RFC[1000]