Coding

Part:BBa_K1740000

Designed by: Yukina Tanimura,Keisuke Sugahara,   Group: iGEM15_KAIT_Japan   (2015-08-20)

Dronpa 145N ORF (E. coli optimized)
This part is the open reading frame of optimized Dronpa 145N to E.coli. Dronpa 145N mutated wild-Dronpa which changed from 145Lys to Asn. Dronpa 145N switched on under 390nm light from a non-fluorescent state into a fluorescent state, and back again under 503nm light. At the fluorescent state, Dronpa 145N forms a tetramer which is combined with other Dronpa 145N. On the other hand, at the non-fluorescent state, Dronpa 145N forms a monomer. This is reversible action, and it is controlled by light. Dronpa 145N is one of the Green Fluorescent Proteins, it is in its on-state, displaying bright green fluorescence.



we expressed Dronpa 145N(BBa_K1740000) in Ecoli(DH5α).
Kekka1.PNG Kekka2.PNG
Figure1 shows the Ecoli that expressed was irradiated with 400nm light. Figure2 show the Dronpa 145N irradiated with 400nm light. We confirmed Dronpa 145N were bright green fluorescence under 400nm light.


We irradiated 400 nm (5 hours) and 500 nm (5 hours) collectively for Dronpa 145K and 145N., we analyzed for fluorescence intensity of Dronpa 145N and Dronpa 145K. Figure3 shows the result of fluorescence intensity analysis.
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Fluorescence intensity of Dronpa 145K was bigger than that of Dronpa 145N in each state. We confirmed that Dronpa’s fluorescence intensity was becoming stronger when we irradiated Dronpa with 400 nm. And it was becoming weaker when we irradiated Dronpa with 500 nm. We proved by specific wavelength.
In addition to this result, we did more experiment, SDS-PAGE, Native-PAGE, Confocal laser scanning microscope (CLSM).


More detail of our result(http://2015.igem.org/Team:KAIT_Japan/Results)
We confirmed this par sequence.(http://2015.igem.org/wiki/images/1/13/20150817_DronpaN_sequence_confirmed_Fw_%281%29.pdf)


Usage and Biology

Sequence and Features

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


[edit]
Categories
//cds/reporter/gfp
//function/reporter/fluorescence
Parameters
colorGreen