Codons were optimized for E. Coli expression and non compatible restriction sites were removed.
This is a Mcherry reporter protein fused to a functional TetR repressor under the control of the arabinose inducible promoter pBAD on the UNS backbone. This part was originally designed to be used as an upstream circuit simulator for WM16_iGEM's circuit control toolbox. The inducible pBad represents an arbitrary circuit with TetRas the product, and as the input to the orthogonal circuit used to modify the transfer function. The mcherry allows for the quantification of the input in the same cell as the quantification of the output (GFP).
Linker and mCherry taken from Synthetic Analog Computation in Living Cells (Daniel et al.). TetR from NCBI NC_011812.1
Kalen Clifton, Christine Gao, Andrew Halleran, Ethan Jones, Likhitha Kolla, Joseph Maniaci, John Marken, John Mitchell, Callan Monette, Adam Reiss
UNS 2 Sequence, from Torella et al., 2013
Torella, J. P., Boehm, C. R., Lienert, F., Chen, J. H., Way, J. C., & Silver, P. A. (2013). Rapid construction of insulated genetic circuits via synthetic sequence-guided isothermal assembly. Nucleic acids research, gkt860.
This is Unique Nucleotide Sequence 2, (UNS 2), from Torella et al., 2013. The William and Mary iGEM team has adopted this as our standard prefix; as such, all of our parts will have this sequence immediately following the BioBrick prefix. We took this measure in order to allow easier Gibson Assembly cloning of our parts. Primer sequences which can be used to clone with the UNS 2/3 standard can be found on our wiki.
UNS 2 was chosen because it works well with UNS 3 and it is in accordance with the BioBrick standard.
UNS 3 Sequence, from Torella et al., 2013
This is Unique Nucleotide Sequence 3, (UNS 3), from Torella et al., 2013. The William and Mary iGEM team has adopted this as our standard prefix; as such, all of our parts will have this sequence immediately following the BioBrick prefix. We took this measure in order to allow easier Gibson Assembly cloning of our parts. Primer sequences which can be used to clone with the UNS 2/3 standard can be found on our wiki. The sequence for this part came from the following paper: Torella, J. P., Boehm, C. R., Lienert, F., Chen, J. H., Way, J. C., & Silver, P. A. (2013). Rapid construction of insulated genetic circuits via synthetic sequence-guided isothermal assembly. Nucleic acids research, gkt860. A huge thanks to all the researchers involved in its original creation!
This UNS sequence was chosen to serve as the 3' primer in our standard because it works well with UNS 2 and it adheres to the BioBrick standards.
RBS (Elowitz 1999) -- defines RBS efficiency
Varies from -6 to +1 region from original sequence to accomodate BioBricks suffix. No secondary structures are formed in the given RBS region. Users should check for secondary structures induced in the RBS by upstream and downstream elements in the +50 to -50 region, as such structures will greatly affect the strength of the RBS. Contact info for this part: Brian Chow
Vinay S Mahajan, Voichita D. Marinescu, Brian Chow, Alexander D Wissner-Gross and Peter Carr IAP, 2003.
Double terminator consisting of BBa_B0010 and BBa_B0012
Sequence and design concerns taken from Synthetic Analog Computation in Living Cells (Daniel et al.)
This is an Mcherry reporter which is designed to be fused to a transcription factor with a linker sequence. This allows for quantification of transcription factors as well as a GFP reporter.
TetR from NCBI NC_011812.1, Linker from Synthetic Analog Computation in Living Cells (Daniel et al.)
TetR protein designed to be used in fusion proteins.
E. Coli codon optimized. Restriction sites removed
Derived from the TE terminator of T7 bacteriophage between Genes 1.3 and 1.4 <genbank>V01146</genbank>.
Transcription terminator for the E.coli RNA polymerase.
Suggested by Sri Kosuri and Drew Endy as a high efficiency terminator. The 5' end cutoff was placed immediately after the TAA stop codon and the 3' end cutoff was placed just prior to the RBS of Gene 1.4 (before AAGGAG). Use anywhere transcription should be stopped when the gene of interest is upstream of this terminator.
Transcriptional terminator consisting of a 64 bp stem-loop.
Conversion of the iGEM parts registry to SBOL2.1