Part:BBa_K4390004
merR expression construct
BBa_K4390004 is a working transcriptional unit which expresses a MerR repressor (BBa_K3470001). It is composed of the constitutive promoter J23100 (BBa_J23100), a ribosome binding site (BBa_B0034), the MerR repressor sequence which has been adapted to an O part for JUMP assembly (BBa_K4390068) and the weak synthetic L2U2H09 Terminator (BBa_K4390001). The MerR sequence is flanked by the N and C fillers (BBa_K4390006 and BBa_K4390008) so that our transcriptional unit works correctly with JUMP assembly. This unit was assembled using JUMP and can be assembled into a JUMP Level 1 vector plasmid for expression in cells.
Usage and Biology
The merR repressor is derived from the mer operon of gram-negative bacteria, the repressor will bind to the PmerR promoter in the absence of mercury ions. We used this transcriptional unit alongside the Hg biosensor (BBa_K346002) so that when we use cell lysate which expresses our transcriptional unit there would be merR present in the lysate. When this cell-free extract is combined with the Hg biosensor it induces transcriptional repression of the linear biosensor when Hg (II) salts are not present. When Hg(II) salts are present the merR binds to the metal ions and remains bound to the linear biosensor and then acts as an activator of 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
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 11
Illegal NheI site found at 34 - 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
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