Part:BBa_K3425051
β-subunit of Qβ replicase with cleavable degradation tag
BBa_K3425051 is the composite part made up of the β-subunit of Qβ replicase, the TEV protease cleave signal sequence and the LAA degradation tag.
Function
BBa_K3425051 was ordered and used as a control construct, which contains the stop codon TAG between the β-subunit and the cleavage signal. The functional construct can be derived from this part by PCR mutagenesis using custom primers which will remove this stop codon.
When this functional construct is expressed in the absence of TEV protease, the β-subunit is relatively quickly degraded due to the presence of LAA degradation tag. However, upon the expression of TEV protease, the degradation tag is cleaved off and the protein is stabilized. This mechanism is part of the system which reduces noise in the biosensor of Uppsala 2020 team.
Assembly compatibility
Note that the sequence is adapted to Type IIS iGEM standard assembly (RFC1000) and therefore lacks the start codon ATG. When assembled using Type IIS standard, the start codon is included in the scar between the RBS element and the coding sequence part. You can find more information about how to clone the parts using this standard in the Type IIS guide of Uppsala 2020 team.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal BamHI site found at 1314
Illegal XhoI site found at 822
Illegal XhoI site found at 988 - 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
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