Part:BBa_K567001

lacI-Ptrc-tRNA(Arg)

The tRNA^Arg is under the control of promoter trc. tRNA^Arg expression is induced by 0.5mM IPTG when the OD600 of the culture reaches 0.3.

Design

In this part we have overexpressed rare tRNA^Arg-AGG in the cell. The rare tRNA can recognize AGG codon on the mRNA.

tRNA^Arg-AGG(BBa_K567001): tRNA^Arg-AGG is over expressed under the control of trc promoter (induced by IPTG).

This rare tRNA^Arg can be charged with Arg by native Arginyl-tRNA Synthetase(ArgRS) in E.coli.

RFP-6AGG(BBa_K567017): we have inserted 6 AGG codons after the start codon ATG in the RFP gene.

Action

When rare tRNA^Arg-AGG is not over-expressed, RFP expression is hindered. When tRNA^Arg-AGG is over-expressed, this tRNA can recognize the AGG codon on the mRNA so a large amount of RFP is produced.

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Result

Fig.1 Confocal Microscope examining RFP expression. RFP has been largely produced in cells overexpressing tRNA^Arg-AGG.

Fig.2 Cells over-expressing tRNA^Arg-AGG emit bright red fluorescence as wild type RFP (first one from the left). Control (first one from the right) exhibits no red fluorescence.

RFP has been largely produced in cells overexpressing tRNA^Arg. No RFP can be observed in cells without rare tRNA overexpression.

We have successfully controlled protein expression by controlling rare tRNA amount.

Reference

Ulrich Deuschlel., et al., Promoters of Escherichia coli: a hierarchy of in vivo strength indicates alternate structures The EMBO Journal vol.5 no. 11 pp.2987-2994, 1986

Related Biobrick

Pbla-Luc-2AGG (BBa_K567004)

Pbla-Luc-4AGG (BBa_K567005)

Pbla-Luc-6AGG (BBa_K567006)

Pbla-Luc-8AGG (BBa_K567007)

PT7-Luc-2AGG (BBa_K567008)

PT7-Luc-4AGG (BBa_K567009)

PT7-Luc-6AGG (BBa_K567019)

PT7-Luc-8AGG (BBa_K567010)

PT7-RFP-6AGG (BBa_K567017)

PT7-Luc-2x4AGG(111bp) (BBa_K567021)

PT7-Luc-2x4AGG(30bp) (BBa_K567026)

PT7-Luc-3x4AGG(30bp+111bp) (BBa_K567027)

Sequence and Features

Assembly Compatibility:

• 10
  INCOMPATIBLE WITH RFC[10]

  Illegal PstI site found at 1845
  
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal PstI site found at 1845
  
• 21
  COMPATIBLE WITH RFC[21]
• 23
  INCOMPATIBLE WITH RFC[23]

  Illegal PstI site found at 1845
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal PstI site found at 1845
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI site found at 2026