Coding

Part:BBa_K2485012:Design

Designed by: Michael Herschl   Group: iGEM17_Stanford-Brown   (2017-09-20)
Revision as of 19:28, 21 September 2017 by Mherschl (Talk | contribs) (→‎Design Notes)


Proteorhodopsin-cPT-RFP


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Unknown
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Unknown
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


Design Notes

A 5x Gly-Ser linker, BBa_J18922, was used to join proteorhodopsin and cPT.

A 6x His tag was also added for potential protein purification.

Source

The green light absorbing protein rhodopsin was sourced from BBa_K773002 and codons were slightly modified to enhance GC content. It is a proton pump used by marine bacteria for generating proton motive force for ATP generation.

The cPT Region of IcsA was sourced from BBa_K2485011 with removed start and stop codons, and the last four nucleotides were modified to match the fusion sequence between cPT and GFP in a plasmid provided by Marcia B. Goldberg, MD which was experimentally shown to unipolarly localize GFP.

FresnoRFP from Atum was used to maximize the distance between the absorption/emission of the fluorescent protein (553/592) and the absorption of the proteorhodopsin (~520).

References