Reporter
MelA

Part:BBa_K274001:Design

Designed by: Shuna Gould   Group: iGEM09_Cambridge   (2009-08-03)

melanin pigment


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 1332
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 1184
    Illegal BsaI.rc site found at 489


Design Notes

Created by PCR

The only changes to the sequence from the plasmid kindly provided by The Massachusetts Institute of Technology and Gregory Stephanopoulos was the removal of two PstI restriction sites. This was done by converting CAG to CAA in the second codon of the site.

MelA characterization contribution

Group: iGEM Concordia 2016 Author of experiment: Kevin Gorjipour

MelA characterization
alt text

MelA was characterized under a constitutive promoter. For the first time ever in iGEM history, polymerization of tyrosine was characterized. The above graph shows the polymerization of tyrosine by melA under a constitutive promoter in DH5 alpha E. coli for 191 20 minutes time points . 2 controls were performed for this experiment, DH5 alpha without the MelA gene and a no cell control. The no cell control was done in order to verify if tyrosine would spontaneously polymerize in solution. The no MelA control was done in order to verify if tyrosine polymerization was, in fact, due to the presence of MelA expression in the cell. The no cell showed a constant OD at 450nm which pointed to us that tyrosine does not spontaneously polymerize. The constitutive MelA producing cells showed a faster increase in OD450 then the “no MelA” control telling us that MelA is polymerization tyrosine.

We improved the characterization of this part by showing that MelA can be used under a constitutive promoter, without inducing toxicity to the cell, to convert tyrosine into Eumelanin.

Source

The melA gene comes from the Rhizobium etli CFN 42 symbiotic plasmid p42d (accession number U80928). The original plasmid was created by Christine Nicole S. Santos and Gregory Stephanopoulos.

References

Christine Nicole S. Santos and Gregory Stephanopoulos Melanin-Based High-Throughput Screen for L-Tyrosine Production in Escherichia coli American Society for Microbiology 2007 74(4) 1190-1197 doi:10.1128/AEM.02448-07 [http://aem.asm.org/cgi/content/abstract/74/4/1190 reference]