Part:BBa_K2036025
Kozak-Snrk2.2-TTADH1-pCyc
One of the Basic Part of genetic circuit in the eukaryotic version of signal filter (BBa_K2036030).
We construct this circuit for further building of whole eukaryotic version of signal filter and for verifying the function of related eukaryotic proteins.
Protein expression
Pichia pastoris are typical Eukaryotic expression host, but expressing plant genes are still a big challenge. So we firstly tested whether our three interest genes can be efficient expressed. We measured the secretion expression of PP2CA, ABF2 and SnRK2.2. We ran a SDS-PAGE of the culture supernatants of induced cells to identify the three recombinant proteins(Fig1). Compared to vector trasnfected cells, 3 kinds of recombinant protein plasmids trasnfected cells all expressed proteins at ~80kD under induction. The shift of protein bands suggested potential glycosylation modification.
Bi-stable function:
We constructed expression plasmid and submitted this part (BBa_K2036030) to the registry. But due to the limited time, its function characterization is still under testing. However, our modeling simulation showed promising switch functions. [http://2016.igem.org/Team:HUST-China/Model/model-euk See more details in our Eukaryote circuit modeling.]
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal BamHI site found at 15
Illegal BamHI site found at 672 - 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
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