Part:BBa_K4335003
ChlamyHgR
We used the Rbcs2 promoter and terminator, and since the Rbcs2 Promotor Rhizobium-derived transcription factor was low, we added the UTR of the heat shock promoter HSP70A Promotor to enhance the promoter and thus the heterologous expression of the Staygold fluorescent protein StayGold, a green fluorescent protein (GFP) from the jellyfish family Endocystis, was chosen as our reporter fluorescent protein because it is orders of magnitude more photostable than any currently available fluorescent protein, and because the excitation and emission wavelengths of Staygold do not overlap with the chloroplast fluorescence of Chlamydomonas reinhardtii itself.
Usage
We inserted ChlamyHgR into plasmids pTX2038 and pTX2040 as marker genes for transformation screening.Result
Plasmid construction
To verify our successful assembly, we designed two primers, Hgy-F and Hgy-R, for PCR verification. The primer location, primer sequence and gel map are shown in the following figure:
mCherry and the position of primer targeting
Electrophoregram of amplification products,M is DNA Marker.
Validation of conversion results
Fluorescence excitation of positive clones transferred into pTX2038 vector compared with wild type mCherry, together with DIC field and chlorophyll excitation as fluorescence control.
Reference
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal NgoMIV site found at 782
Illegal NgoMIV site found at 890 - 1000COMPATIBLE WITH RFC[1000]
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