Uranium Reducing and Detection Device for E. coli
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.
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.
TorA Signal Peptide - Silver Fusion Compatible
Chromate and Uranium Reductase, ChrR30 Silver-fusion compatible
This part comes from Caulobacter crescentus CB15 section 318 of 359 of the complete genome, subsection 7998-8971, which may be found in Genbank at accession number AE005992.1 (Hillson et al., US Patent, Feb. 6 2008). Reference: Nathan J. Hillson, Ping Hu, Gary L. Andersen and Lucy Shapiro. Heavy Metal Biosensor. US Patent Application Publication. US20110117590. Feb. 6 2008. This reference was retrieved from: file:///C:/Users/Owner/Desktop/US20110117590.pdf
The promoter was designated urcA for uranium response in the bacterium Caulobacter. It is activated in the presence of the uranyl cation, a soluble form of uranium (Hillson et al., 2007). The uranyl ion (UO2,2+ ) is the most water-soluble and bioavailable form of uranium and poses the greatest threat to human health. Due to the ease of uranyl ion spread through groundwater systems, most bioremediation strategies attempt to prevent contaminating uranium spread by utilizing microorganisms to reduce the oxidation state of uranium from U(VI), found in uranyl, to less soluble forms of uranium, including U(IV)(Hillson et al., 2007). The urcA promoter is speciﬁc for uranium and has little cross speciﬁcity for nitrate (<400 M), lead (<150 M), cadmium (<48 M), or chromium (<41.6 M). It was induced 27.5-fold under uranium stress, but was not upregulated in response to other heavy metals in the test screen. For this reason, the urcA promoter was selected as a candidate to drive uranium reporter constructs(Hillson et al., 2007). Usually promoters from Caulobacter species will include at least one uranium-specific m_5 motif sequence. The urcA promoter contains two matches to this motif, located 107 and 55 bp upstream of the putative +1 site (Hillson et al., 2007). Reference: Nathan J. Hillson, Ping Hu, Gary L. Andersen and Lucy Shapiro. Caulobacter crescentus as a Whole-Cell Uranium Biosensor. Appl. Environ. Microbiol. 2007, 73(23):7615. DOI: 10.1128/AEM.01566-07. This reference was retrieved from: file:///C:/Users/Owner/Desktop/Appl.%20Environ.%20Microbiol.-2007-Hillson-7615-21.pdf
The sequence complied with RFC-10 standard assembly, so no design considerations had to be considered during the creation of the sequence as a biobrick.
GFPuv comes was originally isolated from the bioluminescent jellyfish Aequorea Victoria (Chin et al.).
Green fluorescent protein (GFP) is a 27 kDa protein of 238 amino acids in length. GFP is intrinsically fluorescent and hence is ideal as a non-invasive marker in living cells. It is used as a cell lineage tracer, reporter of gene expression, a measure of protein-protein interactions, and as an intracellular pH indicator. GFP can function as a protein tag as it tolerates N- and C-terminal fusion to a broad variety of proteins (Chin et al.). Wild-type GFP fluoresces maximally when excited at 395 nm, with a minor peak at 475 nm. Fluorescence emission peaks at 509 nm. It is stable within a range of pH 5.5 to 12. Denaturation occurs at temperatures above 70??C (Chin et al.). Several variants with improved absorption or emission spectra, or increased the fluorescence intensity, have since been developed. Variants also exhibit different thermosensitivities (Chin et al.). As stated before, the maximum excitation spectra of GFPuv is 395nm while its maximum emission spectra is 509nm, which is similar to that of wild-type GFP. Compared to wild-type GFP, GFPuv contains three amino acid substitutions (Phe-99 to Ser, Met-153 to Thr, and Val-163 to Ala [based on the amino acid numbering of wild-type GFP]), none of which alter the chromophore sequence. The main advantage of this variant over wild-type GFP is its large amount of fluorescence when excited by ultra-violet (UV) light at 395nm. GFPuv is 18 times brighter than wild-type GFP under ultra-violet excitation and can easily be detected by the eye without the need for specialized equipment. Furthermore, GFPuv expressed in E. coli is a soluble, fluorescent protein in contrast to wild-type GFP, much of which may be expressed as a nonfluorescent protein in inclusion bodies. Consequently, the GFPuv gene is expressed very efficiently in E. coli (Chin et al.). Reference: Chin et al. Characterization of Green Fluorescent Protein. Retrieved from: http://staff.science.nus.edu.sg/~scilooe/srp_2003/sci_paper/dbs/research%20_paper/ngeow_kao_chin.pdf
The sequence complied with RFC-10 assembly compatibility as a requirement for Utah State University Synthetic Biology class, so no design considerations had to be taken into account during the creation of the sequence as a biobrick.
Conversion of the iGEM parts registry to SBOL2.1