Difference between revisions of "Part:BBa M36921:Design"
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===Source===
 
===Source===
  
−
  The promoter and ZntR regulatory gene of the znt operon are naturally existing in E. coli and were first sequenced and amplified from E. coli DH5a genomic DNA. All of our parts were pre-existing and sourced from the iGEM registry, making only slight modifications (silent mutations) to the original ZntR gene (BBa_M11082)  in order to reduce the base pair lengths of palindromic sequences. DNA2.0 also made modifications on the ZntR gene to optimize the sequence for E. coli.  
+
  The promoter and ZntR regulatory gene of the znt operon are naturally existing in E. coli and were first sequenced and amplified from E. coli DH5a genomic DNA.  
 +
 
 +
All of our parts were pre-existing and sourced from the iGEM registry, making only slight modifications (silent mutations) to the original ZntR gene (BBa_M11082)  in order to reduce the base pair lengths of palindromic sequences.  
 +
 
 +
DNA2.0 also made modifications on the ZntR gene to optimize the sequence for E. coli.  
  
 
===References===
 
===References===
  
 
Chaudhari, Aparna, and P. Babu. "Development of a Broad Spectrum Fluorescent Heavy Metal Bacterial Biosensor." PubFacts. PubMed, 7 Mar. 2012. Web. 20 Nov. 2014. <http://www.pubfacts.com/fulltext_frame.php?PMID=23070906&title=Development%20of%20a%20broad-spectrum%20fluorescent%20heavy%20metal%20bacterial%20biosensor>.
 
Chaudhari, Aparna, and P. Babu. "Development of a Broad Spectrum Fluorescent Heavy Metal Bacterial Biosensor." PubFacts. PubMed, 7 Mar. 2012. Web. 20 Nov. 2014. <http://www.pubfacts.com/fulltext_frame.php?PMID=23070906&title=Development%20of%20a%20broad-spectrum%20fluorescent%20heavy%20metal%20bacterial%20biosensor>.

Revision as of 20:48, 4 December 2014

Zinc Sensor


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal PstI site found at 194
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 7
    Illegal NheI site found at 30
    Illegal PstI site found at 194
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 190
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal PstI site found at 194
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal PstI site found at 194
  • 1000
    COMPATIBLE WITH RFC[1000]


Design Notes

Our genetic construct consists of a constitutive promoter (BBa_J23108) , a high-affinity ribosome binding site (BBa_J61101) , an optimized sequence of a ZntR regulatory gene (BBa_M36919) that controls the expression of the chromosomal zinc resistance operon znt , a transcriptional terminator consisting of a 64 base pair stem-loop (BBa_B0010) , and a znt promoter sequence with a palindromic binding site for ZntR (BBa_K190016) .

A promoter of medium strength (variant RFP 1303 au) was chosen because we wanted to not put a high translational burden on our cells while still being able to generate enough PoPS signal to guarantee expression of our selected Comet fluorescence output.

This system will be placed into a Comet plasmid with a high-copy origin of replication and Kanamycin resistance as its selection marker.

Source

The promoter and ZntR regulatory gene of the znt operon are naturally existing in E. coli and were first sequenced and amplified from E. coli DH5a genomic DNA.

All of our parts were pre-existing and sourced from the iGEM registry, making only slight modifications (silent mutations) to the original ZntR gene (BBa_M11082) in order to reduce the base pair lengths of palindromic sequences.

DNA2.0 also made modifications on the ZntR gene to optimize the sequence for E. coli.

References

Chaudhari, Aparna, and P. Babu. "Development of a Broad Spectrum Fluorescent Heavy Metal Bacterial Biosensor." PubFacts. PubMed, 7 Mar. 2012. Web. 20 Nov. 2014. <http://www.pubfacts.com/fulltext_frame.php?PMID=23070906&title=Development%20of%20a%20broad-spectrum%20fluorescent%20heavy%20metal%20bacterial%20biosensor>.